171. "Ionic strength effect on the thermal unfolding of
a-spectrin peptides" D. Lusitani, N. Menhart, T. A. Keiderling, L. W.-M. Fung, Biochemistry, 37, 16546-16554 (1998)172. "Effect of resolution enhancement on secondary structure analysis using Fourier Transform IR spectra of proteins in H2O", Sungsool Wi, P. Pancoska, T. A. Keiderling Biospectroscopy 4, 93-106 (1998)
173. "The regiochemistry of reduction of mono-ethyl fumarate and maleate using a ruthenium BINAP catalyst" M. Shaharuzzamam, J. Braddock-Wilking, J. S. Chickos, C. N. Tam, R. A. G. D. Silva, T. A. Keiderling Tetrahedron Asymmetry 9, 1111-1114 (1998)
177. "Novel matrix descriptor for secondary structure segments in proteins: Demonstration of predictability from circular dichroism spectra," P. Pancoska, V. Janota, T. A. Keiderling, Analytical Biochemistry 267, 72-83 (1999)
Novel Matrix Descriptor for Secondary Structure Segments in Proteins. Demonstration of Predictability from Circular Dichroism Spectra
Petr Pancoska*a,b,c, Vit Janotab,c, Timothy A. Keiderlinga*
a Department of Chemistry, University of Illinois at Chicago, 845 W. Taylor Street, M/C 111, Chicago, Il 60607-7061, U.S.A.
b
Center for Discrete Mathematics, Theoretical Computer Science and Applications, Malostranske namesti 25, 118 00 Praha 1, Czech Republicc
Department of Chemical Physics, Faculty of Mathematics and Physics, Charles University, Ke Karlovu 3, 121 16 Praha 2, Czech Republic
Abstract
An extension to standard protein secondary structure predictions using optical spectra is demonstrated that encompasses the number and average lengths of segments of uniform secondary structure in the sequence. The connectivity and numbers of segments can be described by a matrix descriptor [sij] (i,j representing segment types such as helix and
b -sheet strands). Independent knowledge of the fractional concentration of each secondary structure type and of the total number of residues in the protein then with [sij] yields the average segment length of each type. The physical background for prediction of this extended structural descriptor from spectral data is summarized, rules for its generation from reference x-ray structures are defined and formal variants of its form are discussed. Using a novel neural network approach to analyze a training set of electronic circular dichroism (ECD) and vibrational circular dichroism (VCD) spectra for 28 proteins, matrix descriptors encompassing helix, sheet and other forms are predicted. The results show that the matrix descriptor can be predicted to an accuracy comparable to that of conventionally predicted average fractional secondary structures. In this respect the ECD predictions of [sij] were significantly more accurate than the VCD ones which may result from the longer range length dependence of the ECD bandshape and intensity. Summary results for a parallel analysis using Fourier transform Infrared (FTIR) spectra indicate somewhat lower reliability than for VCD.
178. " Vibrational Circular Dichroism Spectroscopy of Selected Oligopeptide Conformations" Timothy A. Keiderling, R. A. G. D. Silva, Gorm Yoder, and Rina K. Dukor, Bioorganic and Medicinal Chemistry 7, 133-141 (1999)
Vibrational Circular Dichroism Spectroscopy of
Selected Oligopeptide Conformations
Timothy A. Keiderling*, R. A. G. D. Silva, Gorm Yoder, Rina K. Dukora
Department of Chemistry, University of Illinois at Chicago,
845 W. Taylor St. (m/c 111), Chicago, IL 60607-7061 USA
Abstract
Vibrational circular dichroism (VCD) has been shown to be a useful technique for characterization of the qualitative secondary structure type for linear polypeptides and oligopeptides. A brief review of characteristic spectral responses and applications is given. Since VCD is dependent on relatively short range interactions, it detects residual structure in such oligomers even if long range order is lost. VCD studies presented here for Lys oligomers as well as Lys and Glu polymers as a function of length, salt added and temperature, confirm residual local order in these "random coils". Comparison to results with Pro oligomers, supports an interpretation that these extended structures have a left handed twist conformation. The "coil" VCD is shown to be significantly reduced in intensity by temperature increase and by decrease in peptide length. By contrast, for partially
a-helical Ac-(AAKAA)3GY-NH2 oligomers, the spectrum changes to the high temperature Lysn shape on heating, first losing then gaining intensity, indicating an equilibrium shift between structured states, from helix to coil (locally ordered) forms. VCD is shown to be a useful technique for monitoring local order in otherwise random coil structures.
179. "Novel use of a static modification of two-dimensional correlation analysis. I. Comparison of the secondary structure sensitivity of electronic circular dichroism, FT-IR, and Raman spectra or proteins" P. Pancoska, J. Kubelka, T. A. Keiderling, Applied Spectroscopy. 53, 655-665 (1999)
180. "Novel use of a static modification of two-dimensional correlation analysis. II. Heterospectral correlations of protein Raman, FT-IR, and circular dichroism spectra" , J. Kubelka, P. Pancoska, T. A. Keiderling, Applied Spectroscopy, 53, 666-671 (1999)
181. "
b -Hairpin stabilization in a 28-residue peptide derived from the b -subunit sequence of human chorionic gonadotropin hormone" R. A. G. D. Silva, Simon A. Sherman, T. A. Keiderling, Biopolymers 50, 413-423 (1999)b
-Hairpin Stabilization in a 28-residue PeptideDerived from the -subunit Sequence of
Human Chorionic Gonadotropin Hormone.
R. A. G. D. Silvaa, Simon A. Shermanb, Timothy A. Keiderlinga *
a Department of Chemistry (M\C 111)
University of Illinois at Chicago, 845 W. Taylor St.,
Chicago IL 60607-7061 USA.
b
Eppley Institute for Research in Cancer and Allied Diseases,986805 Nebraska Medical Center,
Omaha, NE 68189 USA.
Abstract
The b-subunit of human chorionic gonadotropin (hCG) hormone, which is believed to be related to certain types of cancer, contains three hairpin-like fragments. To investigate the role of b-hairpin formation in the early stages of the hCGb folding, a 28-residue peptide with the sequence RDVRFESIRLPGSPRGVNPVVSYAVALS, corresponding to the H3-b hairpin fragment (residues 60-87) of the hCG subunit, was studied under various conditions using three optical spectroscopic methods: Fourier transform infrared spectroscopy, electronic and vibrational circular dichroism. Environmental conditions are critical factors for formation of secondary structure in this peptide. TFE : H2O mixed solvents induced helical formation. Formation of b-structure in this peptide, which may be related to the native b-hairpin formation in the intact hormone, was found to be induced only under conditions such as high concentration, high temperature, and the presence of non-micellar SDS concentrations. These findings support a protein folding mechanism for the hCG subunit in which an initial hydrophobic collapse, that increases inter-molecular interactions in hCGb is needed to induce the H3-b hairpin formation.
187. "Enhanced prediction accuracy of protein secondary structure using hydrogen exchange Fourier Transform Infrared spectroscopy" B. I. Baello, P. Pancoska, T. A. Keiderling, Analytical Biochemistry 280, 46-57 (2000)
188. "Simulations of oligopeptide vibrational CD. Effects of isotopic labeling." Petr Bour, Jan Kubelka,T. A. Keiderling Biopolymers 53, 380-395 (2000)
Simulations of Oligopeptide Vibrational Circular Dichroism. Effects of Isotopic Labelling.
Petr Bour,a Gangani Silvab and Timothy A. Keiderlingb
a
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo nam 2, 16610, Praha 6, Czech Republic,bour@uochb.cas.cz, fax (420-2)-2431-0503
b
Department of Chemistry (M/C 111), University of Illinois at Chicago,845 W. Taylor Street, Chicago, Illinois 60607-7061, U.S.A.,
tak@uic.edu, fax 312-996-0431
Abstract
Simulated infrared (IR) absorption and vibrational circular dichroism (VCD) spectra of four alanine-based octapeptides, each having its main chain constrained to a different secondary structure conformation, were analyzed and compared with experimental results. The octapeptide simulations were based on transfer of property tensors from a series of ab initio calculations for a short L-alanine based segment containing 3 peptide bonds with relative
f,y angles fixed as appropriate for a-helix, 3-10 helix, ProII helix and b-sheet like strand. If one allows for frequency correction, the octapeptide results obtained are consistent with experimental observations for VCD spectra of polypeptides and oligopeptides in established conformations. Predicted VCD changes upon deuteration of the peptide and side-chains as well as 13C isotopic labelling in the peptide chain were compared to experimental results. Apparently, such a combination of modeling and experiment for labeled compounds leads to enhanced resolution and applicability of the VCD spectroscopy.
193. "Site specific conformational determination in thermal unfolding studies of helical peptides using vibrational circular dichroism with isotopic substitution" R. A. G. D. Silva, Jan Kubelka, Petr Bour, Sean M. Decatur, Timothy A. Keiderling, Proceedings of the National Academy of Sciences (USA) 97, 8318-8323 (2000)
Site-Specific Conformational Determination in Thermal Unfolding Studies of Helical Peptides using Vibrational Circular Dichroism with Isotopic Substitution.
R. A. G. D. Silva,1 Jan Kubelka,1 Petr Bour,3 Sean Decatur,2
and Timothy A. Keiderling1*
1Department of Chemistry, University of Illinois at Chicago,
845 W. Taylor St. (M/C 111), Chicago IL 60607-7061, USA;
2Department of Chemistry, Mt. Holyoke College, South Hadley, MA 01075 USA;
3Institute of Organic Chemistry and Biochemistry, Academy of Science,
Prague, Czech Republic
*To whom correspondence should be addressed at UIC. E-mail: tak@uic.edu
Abstract
Understanding of the detailed mechanism of protein folding requires dynamic, site-specific stereochemical information. For a series of blocked, alanine-rich, 20 residue peptides that form stable
a-helices at low temperature in aqueous solution, we used isotopic labeling and vibrational circular dichroism (VCD) to demonstrate that the a-helix unwinds from the ends in an non-cooperative manner with increasing temperature. The C-terminal is shown to be frayed at 5 C. Ab initio level theoretical simulations of the IR and VCD bandshapes are used to analyze the spectra and to confirm the conformation of the labeled and unlabeled components. The intrinsic time characteristic of infrared-based spectroscopies allows evaluation of the distribution of populations in rapid equilibrium as the peptide unfolds, and isotopic labels along with local stereochemical sensitivity of VCD allow determination of the specific sites of unfolding. The VCD signals associated with the labeled residues are amplified by coupling to the non-labeled parts of the molecule. Thus relatively small amounts of label become detectable and stereochemically interpretable in a moderately large peptides in this first report of site-specific peptide VCD.
194. "Folding studies on the human chorionic gonadotropin -subunit using optical spectroscopy of peptide fragments" R. A. G. D. Silva, S. A. Sherman, F. Perini, E. Bedows, T. A. Keiderling, Journal of the American Chemical Society, 122, 8623-8630 (2000)
Folding Studies of the
b-subunit of the Human Chorionic GonadotropinUsing Optical Spectroscopy of Peptide Fragments
R. A. Gangani D. Silvaa, Simon A. Shermanb, Fulvio Perinid, Elliott Bedowsb,c,d and Timothy A. Keiderlinga*
a
Department of Chemistry (M/C 111), University of Illinois at Chicago, 845 W. Taylor St., Chicago, IL 60607-7061 USA.b
Eppley Institute for Research in Cancer and Allied Diseases, cThe Department of Biochemistry and Molecular Biology, dThe Department of Pharmacology, 986805 Nebraska Medical Center, Omaha, NE 68189-6805 USA.Abstract
Conformational preferences for the peptides, SRPINATLAVEKEGSPVS- ITVNTTISA (H1) and APTMTRVLQGVLPALPQVVCNYR (H2), corresponding to the 9-35 and 38-60 regions of the
b subunit of human chorionic gonadotropin (hCG), a glycoprotein hormone, were studied by Fourier transform infrared spectroscopy (FTIR), vibrational circular dichroism (VCD), and electronic circular dichroism (ECD) in various environments. These peptides correspond to the H1- and H2-b hairpins of the native-state hCGb subunit. The H1 peptide adopts a -structure in water and in environments that normally induce -helix formation, such as mixed TFE : H2O solvent or micellar concentrations of SDS. By contrast, the H2 peptide spectra are consistent with a significant fraction of the residues being in a poly proline II (PII) like structure or in a partially helical conformation in all of the environments studied. A third peptide, H3, corresponding to the 60-87 region of the hCGb, was studied previously and it switches its conformation depending on both solvent and concentration. Taken together, the data suggest that hCGb folds using the following mechanism. First, the H1 region forms a b-hairpin followed by hydrophobic collapse with the H3 region that induces formation of the H3 b hairpin. The folding intermediate that brings these two hairpins H1 and H3 together would then lead to the formation of the H2 hairpin loop.
196. "The anomalous infrared amide I intensity distribution in 13C isotopically labeled peptide -sheets comes from extended, multiple stranded structures. An Ab Initio study." Jan Kubelka and T. A. Keiderling , Journal of the American Chemical Society. . 123, 6142-6150 (2001)
Erratum: J. Am. Chem. Soc. 123, 8163 (2001)
Thermal Unfolding of Ribonuclease A in Phosphate at Neutral pH: Deviations from the Two State Model
Simona D. Stelea, Petr Pancoska, Albert S. Benight
and Timothy A. Keiderling*
Department of Chemistry, University of Illinois at Chicago,
845 W. Taylor St., Chicago, IL 60607-7061 USA
* To whom the correspondence should be addressed: Telephone: (312) 996-3156;
Fax: (312) 996-0431; E-mail: tak@uic.edu
ABSTRACT
The thermal denaturation of ribonuclease A (RNase A)1 in the presence of phosphate at neutral pH was studied by differential scanning calorimetry (DSC) and a combination of optical spectroscopic techniques in order to probe the existence of intermediate states. Fourier Transform Infrared (FTIR) spectra of the amide I' band and far-uv circular dichroism (CD) spectra were used to monitor changes in the secondary structure. Changes in the tertiary structure were monitored by near-uv CD. Spectral bandshape changes with change in temperature were analyzed using factor analysis. The global unfolding curves obtained from DSC confirmed that structural changes occur in the molecule prior to the main thermal denaturation transition. The analysis of the far-uv CD and FTIR spectra showed that these lower temperature induced modifications occur in the secondary structure. No pre-transition changes in the tertiary structure (near-uv CD) were observed. The initial changes observed in far-uv CD were attributed to the fraying of the helical segments, which would explain the loss of spectral intensity with almost no modification of spectral bandshape. Separate analyses of different regions of the FTIR amide I' band suggest that, in addition to -helix, part of the pre-transitional change also occurs in the -strands.
199. "Polarization Modulation Fourier Transform Infrared Spectroscopy with Digital Signal Processing: Comparison of Vibrational Circular Dichroism Methods." Jovencio Hilario, David Drapcho, Raul Curbelo, Timothy A. Keiderling, Applied Spectroscopy 55, 1435-1447 (2001)--accelerated paper
Polarization Modulation Fourier Transform Infrared Spectroscopy with Digital Signal Processing. Comparison of Vibrational Circular Dichroism Methods.
Jovencio Hilario1, David Drapcho2, Raul Curbelo2, and Timothy A. Keiderling1
1 Department of Chemistry, University of Illinois at Chicago
845 W. Taylor Street, Chicago. IL 60607-7061
2
Bio-Rad Laboratories, Digilab Division237 Putnam Avenue, Cambridge, MA 02139
Abstract
Digital Signal Processing (DSP) has been implemented in a step-scan FTIR spectrometer in a modification that enables processing of high frequency polarization modulation signals. In this work, direct comparison is made between vibrational circular dichroism (VCD) spectra measured on the same instrument, with the same samples, under the same conditions using this new DSP method and a conventional rapid-scan technique (employing a lock-in amplifier for demodulation). In this initial test, both techniques generated high quality VCD for solution phase, rigid chiral molecules such as -pinene and camphor. Noise and reproducibility of known spectral features as well as enhancing signal measurability and discrimination were used as criteria for the selection of optimal DSP measurement parameters. Both DSP and rapid-scan VCD methods produced reasonable spectra for biologically relevant molecules such as poly--benzyl-L-glutamate, poly-L-proline, and duplex RNA homopolymer. In most cases the DSP method had a slight signal-to-noise advantage over the rapid-scan measurement, but the final results did depend on the details of the data collection and the phase correction methods inherent in both methods.
200. "Differentiation of -sheet forming structures: ab initio-based simulations of IR absorption and vibrational CD for model peptide and protein -sheets" Jan Kubelka and T. A. Keiderling, Journal of the American Chemical Society. 123, 12048-12058 (2001).
Differentiation of
b-Sheet Forming Structures: ab Initio Based Simulations of IR Absorption and Vibrational CD for Model Peptide and Protein b-SheetsJan Kubelka and Timothy A. Keiderling*
Department of Chemistry (M/C 111), University of Illinois at Chicago,
845 W. Taylor Street, room 4500, Chicago IL 60607-7061, USA
Abstract
Ab initio quantum mechanical computations of force fields (FF) and atomic polar and axial tensors (APT and AAT) were carried out for Ac-A-A-NH-CH3 in single, double and triple strand -sheet like conformations. Models with , , and angles constrained to values appropriate for planar antiparallel and parallel as well as coiled antiparallel (two-stranded) and twisted antiparallel and parallel sheets were computed. The FF, APT and AAT values were transferred to corresponding larger oligopeptide -sheet structures of up to 5 strands of 8 residues each, and their respective IR and vibrational circular dichroism (VCD) spectra were simulated. The antiparallel planar models in a multiple-stranded assembly give a unique IR amide I spectrum with a high intensity low frequency component, but have very weak negative amide I VCD, both closely reflecting experimental patterns seen in aggregated structures. Parallel and twisted -sheet structures do not develop a highly split amide I, making their IR spectra non-discriminating. These have more intense, predominantly negative VCD, reflecting typical experimental data for -sheet containing proteins, but the detailed bandshapes vary considerably. Twist in the antiparallel -sheet structure leads to a significant increase in VCD intensity, while the parallel structure was not as dramatically affected by the twist. Overall predicted VCD intensity is quite weak, which can explain the high variation seen experimentally in -forming peptides and proteins. Even larger variation was predicted in the amide II VCD, which had added complications due to non-hydrogen bonded residues on the edges.
Ab Initio Calculation of Amide Carbonyl Stretch Vibrational Frequencies in Solution with Modified Basis Sets. I. N-Methyl Acetamide.
Jan Kubelka and Timothy A. Keiderling*
Department of Chemistry (M/C 111), University of Illinois at Chicago
845 W. Taylor Street, Chicago IL 60607-7061, USA.
Abstract
Density functional theory DFT(BPW91) level calculations with modified 6-31G(d) basis sets are tested for a small amide, N-methyl acetamide (NMA), as an efficient way for calculating amide I and amide II frequencies that are directly comparable to those commonly measured in solution. The calculational results are compared to experimentally measured FTIR spectra in gas and solution phases. The 6-31G(d) basis set at the DFT level yields vibrational frequencies that have the best agreement with the gas phase experiment, as compared to amide I and II frequencies calculated with the same basis at the HF, CASSCF, MP2, QCISD and CCD levels. The DFT(BPW91)/6-31G(d) level calculation for the NMA
× 3H2O hydrogen bonded complex with an Onsager or CPCM reaction field yields amide I, II and III frequencies comparable to the experiment in aqueous solution. The amide I and, to a smaller degree, amide II frequencies are found to be sensitive to the exponent of the d-function in the basis set. Use of more diffuse (smaller exponent) d-functions in the 6-31G(d) basis set results in a calculated amide I frequency closer to the solution experimental values. Such modified, relatively small basis sets may provide a computationally efficient means of approximating the solvent effects on amide vibrational frequencies.
202. "Chirality in peptide vibrations. Ab initio computational studies of length, solvation, hydrogen bond, dipole coupling and isotope effects on vibrational CD. " Jan Kubelka, Petr Bour, R. A. Gangani D. Silva, Sean M. Decatur, Timothy A. Keiderling, ACS Symposium Series 810, ["Chirality: Physical Chemistry," (Ed. Janice Hicks) American Chemical Society, Washington, DC ](2002), pp. 50-64
203. "Spectroscopic Characterization of Selected -Sheet Hairpin Models", J. Hilario, J. Kubelka, F. A. Syud, S. H. Gellman, and T. A. Keiderling. Biopolymers (Biospectroscopy) 67: 233-236 (2002)
Spectroscopic Characterization of Selected Beta-Sheet Hairpin and Three-Stranded Beta-Sheet Models
J. Hilarioa, J. Kubelkaa, F. Syudb, S. Gellmanb, and T. A. Keiderlinga*
a
Department of Chemistry, University of Illinois at Chicago, 845 W. Taylor St., Chicago IL 60607-7061, USAb
Department of Chemistry, University of Wisconsin Madison, Madison, WIAbstract
The importance of understanding structure, stability, formation and design of -sheets has grown beyond their obvious role as the second most populated protein secondary structure type. New interest is based on their role in protein recognition and diseases resulting in fibril formation. Infrared and vibrational circular dichroism (IR and VCD) spectra were used in conjunction with electronic CD (ECD) to characterize a model two-stranded -hairpin and compare it to a related cyclic two-stranded and a three stranded -sheet model stabilized by a D-Pro-Gly (type II'). Vibrational spectra were compared to ab initio based spectral simulations based on the cyclic peptide structure to determine specific assignments or the dominant features. Simulations show reasonable agreement with experimental amide I spectra for model -hairpins.
204. " Discrimination between peptide 310- and -helices. Theoretical analysis of the impact of -methyl substitution on experimental spectra " Jan Kubelka, R. A. Gangani D. Silva, and T. A. Keiderling, Journal of the American Chemical Society, 124, 5325-5332 (2002)
Spectroscopic Discrimination between Peptide 310- and
a-Helices. Study of the Impact of a-Methyl SubstitutionJan Kubelka, R. A. Gangani D. Silva and Timothy A. Keiderling*
Contribution from the Department of Chemistry (M/C 111),University of Illinois at Chicago,
845 West Taylor Street,Chicago, Illinois 60607-7061
Abstract: Detailed spectral simulations based on ab initio DFT (density functional theory) computations of the amide I and II infrared (IR) and vibrational circular dichroism (VCD) spectra for Ac-(Ala)4-NH2, Ac-(Aib-Ala)2-NH2, and Ac-(Aib)4-NH2 constrained to 310- and -helical conformations are presented. Parameters from these ab initio calculations are transferred onto corresponding larger oligopeptides to simulate the spectra for dodecamers. The differences between conformations and for different Aib substitution patterns within a conformation are reflected in observable spectral patterns where data are available. Simulated IR spectra show small frequency shifts in the amide I maxima between 310- and -helices, but the same magnitude shifts occur within one conformation upon Aib substitution. Thus, from a computational basis, the frequency of the amide I maximum does not discriminate between the 310- or -helical conformations. Calculated VCD band shapes for 310-helices showed more significant changes in amplitude, with change in the fraction of Aib, than those for -helices. Generally, with increasing Aib content the overall amide I VCD intensity becomes weaker and the amide I couplet becomes more conservative, while the amide II VCD is less affected. Although the detailed bandshape is shown to be sensitive to -Me substitution, the basic pattern of amide I and II relative VCD intensities still differs between - and 310-helices and, as a consequence, successfully discriminates between them.
205.? "Ab Initio Quantum Mechanical Models of Peptide Helices and their Vibrational Spectra" Petr Bour, Jan Kubelka and T. A. Keiderling, Biopolymers 65, 45-59 (2002)
Ab Initio
Quantum Mechanical Models of Peptide Helices and Their Vibrational SpectraPetr BouR,a Jan Kubelka,b Timothy A. Keiderlingb*
aInstitute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo nám. 2, 16610, Praha 6, Czech Republic,
and
b
Department of Chemistry (M/C 111), University of Illinois at Chicago, 845 W. Taylor Street, Chicago, Illinois 60607-7061, U.S.A.ABSTRACT
Structural parameters for standard peptide helices (a -, 310, 31 left-handed) were fully ab initio optimized for Ac-(L-Ala)9-NHMe and for Ac-(L-Pro)9-NHMe (PLP I and PLP II forms), in order to better understand the relative stability and minimum energy geometries of these conformers and the dependence of the infrared (IR) absorption and vibrational circular dichroism (VCD) spectra on detailed variation in these conformations. Only the 310-helical Ala-based conformation was stable in vacuum for this deca-amide structure, but both Pro-based conformers minimized successfully. Inclusion of solvent effects, by use of the COSMO continuum model, enabled ab initio optimizations (at the DFT/B3LYP/SV(P) level) without any constraints for the a - and 310-helical Ala-based peptides as well as the two Pro-based peptides. The geometries obtained compare well with peptide chain torsion angles and hydrogen bond distances found for these secondary structure types in X-ray structures of peptides and proteins. For the simulation of VCD spectra, force field and intensity response tensors were obtained ab initio for the complete Ala-based peptides in vacuum, but constrained to the COSMO optimized torsional angles, due to limitations of the solvent model. Resultant spectral patterns reproduce well many aspects of the experimental spectra and capture the differences observed for these various helical types.
206. "Pretransitional Structural Changes in the Thermal Denaturation of Ribonuclease S and S protein"" Simona D. Stelea, T. A. Keiderling, Biophysical Journal 83, 2259-2269 (2002)
Pre-transitional structural changes in the thermal denaturation of ribonuclease S and S protein
Simona D. Stelea and Timothy A. Keiderling*
Department of Chemistry, University of Illinois at Chicago,
845 W. Taylor St., Chicago, IL 60607-7061 USA
ABSTRACT
Two mechanisms have been proposed for the thermal unfolding of ribonuclease S (RNase S). The first is a sequential partial unfolding of the S peptide/S protein complex followed by dissociation, while the second is a concerted denaturation/dissociation. The thermal denaturation of ribonuclease S and its fragment, the S protein, were followed with circular dichroism and infrared spectra. These spectra were analyzed by the principal component method of factor analysis. The use of multiple spectral techniques and of factor analysis monitored different aspects of the denaturation simultaneously. The unfolding pathway was compared to that of the parent enzyme ribonuclease A (RNase A), and a model was devised to assess the importance of the dissociation in the unfolding. The unfolding patterns obtained from the melting curves of each protein imply the existence of multiple intermediate states and/or processes. Our data provide evidence that the pre-transition in the unfolding of ribonuclease S is due to partial unfolding of the S protein/S peptide complex and that the dissociation occurs at higher temperature. Our observations are consistent with a sequential denaturation mechanism in which at least one partial unfolding step comes before the main conformational transition which is instead a concerted, final unfolding/dissociation step.
209. " Discriminating 310- from -helices. Vibrational and electronic CD and IR Absorption study of related Aib-contining oligopeptides " R. A. Gangani D. Silva, Sritana Yasui, Jan Kubelka, Fernando Formaggio, Marco Crisma, Claudio Toniolo, and Timothy A. Keiderling, Biopolymers 65, 229-243 (2002)
Discriminating 310- from
a-Helices:A Vibrational and Electronic CD and IR Absorption Study of Related Aib-Containing Oligopeptides
R. A. Gangani D. Silvaa, Sritana C. Yasuia, Jan Kubelkaa, Fernando Formaggiob,
Marco Crismab, Claudio Toniolob and Timothy A. Keiderlinga,*
a
Department of Chemistry (M/C 111), University of Illinois at Chicago,845 W. Taylor Street, Chicago, IL 60607-7061, USA
b
Biopolymer Research Center, CNR, Department of Organic Chemistry,University of Padova, 35131 Padova, Italy
Abstract: Model peptides based on -(Aib-Ala)n-, and (Aib)n-Leu-(Aib)2 sequences, which have varying amounts of 310-helical character, were studied by use of vibrational and electronic circular dichroism (VCD and ECD) and Fourier transform infrared (FTIR) absorption spectroscopies, to test the correlation of spectral response and conformation. The data indicate that these peptides, starting from a length of about four to six residues, predominantly adopt a 310-helical conformation at room temperature. The longest model peptides, depending on the series, may evidence some -helical contribution to the spectra, while the shorter ones, with less than six residues, have much less order. The IR absorption spectra (as supported by theory) showed only small frequency changes between 310- and -helices. By contrast, solvent effects are a source of much bigger perturbations. The ECD results show that the intensity ratio for the ~222 nm to ~208 nm bands, while useful for distinguishing between these two helical types in some sequences, may have a narrower range of application than VCD. However, the VCD data presented here continue to support the proposed discrimination between - and 310-helices based on qualitative amide I and II bandshape differences. The present study shows the intensity ratios of the 310-helical amide I (peak-to-peak) to its amide II VCD to be the order of 1-2 and useful for discriminating them from -helices, whose amide I dominates the amide II in intensity. This qualitative result is experimentally independent of the amount of Me substituted residues in the sequence. These experimental VCD results are consistent in detail with theoretical spectral simulations for Ac-(Ala)8-NH2, Ac-(Aib-Ala)4-NH2, and Ac-(Aib)8-NH2 in 310- and -helical conformations.
210. " Partial optimization of molecular geometry in normal coordinates and use as a tool for simulation of vibrational spectra" Petr Bour, T. A. Keiderling, Journal of Chemical Physics 117, 4126-4132 (2002).
Partial optimization of molecular geometry in normal coordinates and use as a tool for simulation of vibrational spectra
Petr Boura
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo nám. 2, 16610, Praha 6, Czech Republic, bour@uochb.cas.cz
Timothy. A. Keiderling
Department of Chemistry (M/C 111), University of Illinois at Chicago, 845 W. Taylor Street, Chicago, Illinois 60607-7061, U.S.A., tak@uic.edu
Abstract
A normal mode coordinate-based molecular optimization algorithm was implemented and its performance tested against other optimization techniques. In certain cases the method was found to be computationally simpler and numerically more stable than the optimization algorithms based on Cartesian or internal valence coordinates. The usual redundant/internal coordinate scheme provided the fastest convergence for compact covalently bond molecules, while the normal mode method was found to be more suitable for more weakly-bond molecular complexes. For constrained optimizations use of the normal coordinates allows one to naturally separate the lower-energy modes from those more typically studied with vibrational spectroscopy. Thus it provides an appropriate tool for simulations of IR and Raman spectra of larger molecules and complex systems when specific conformations are desired.
In Press
Submitted for Publication
211. "Spectroscopic Studies of Structural Changes in Two -Sheet Forming Peptides Show an Ensemble of Structures That Unfold Non-Cooperatively" Serguei V. Kuznetsov, Jovencio Hilario, T. A. Keiderling, Anjum Ansari, Biochemistry (submitted)
212. " Optical spectroscopic investigations of model -sheet hairpins in aqueous solution" Jovencio Hilario, Jan Kubelka, T. A. Keiderling, Journal of the American Chemical Society (to be submitted)
This is a review, 2002, vol.62:
Chapter for Advances in Protein Chemistry
Ed. George Rose
UNFOLDED PEPTIDES AND PROTEINS STUDIED WITH INFRARED ABSORPTION AND VIBRATIONAL CIRCULAR DICHROISM SPECTRA
TIMOTHY A. KEIDERLING and QI XU
Department of Chemistry, University of Illinois at Chicago,
845 W. Taylor St. (m/c 111), Chicago, IL 60607-7061 USA
Abstract
Methods of using infrared absorbance (IR) and vibrational circular dichroism (VCD) spectroscopy for the study of unfolded proteins and peptides are presented. Methods for assigning conformation from spectra based on empirical correlations with model spectra and theoretical modeling of peptide oligomer spectra are discussed. Examples of peptides and protein spectra corresponding to unfolded structures and unfolding transitions are described.
This is abstract for Gangani Thesis 2000:
ABSTRACT
Human Chorionic Gonadotropin (hCG) is a hormone important in maintaining early stages of pregnancy. This is also used as a marker for detecting certain types of cancers. Here we report folding studies of the three hairpin loops of the -subunit of hCG using complementary optical spectroscopic techniques: electronic circular dichroism (ECD), vibrational circular dichroism (VCD) and infrared absorbance (IR). Our studies, which exposed the peptides to different environments, brought out their hidden intrinsic propensities towards particular secondary structures. Accounting for the clear variance in propensities of these peptides, a novel folding mechanism was proposed for hCG subunit folding; according to which the formation of first hairpin (H1) hydrophobically collapses with the second sequence (H3), facilitating formation of that hairpin and finally locking the third sequence (H2) into a loop.
The thesis next focuses on the most common secondary structural type in proteins, the -helix. Segment specific stability of a selected -helix was analyzed by using five model peptides, Ac-(AAAAK)3AAAAY-NH2 with 13C labels introduced onto selected consecutive Ala residues at the backbone carbonyls. VCD together with IR spectra for the amide I region of these peptides in D2O at 5 oC showed a difference in structure for the labeled segments of the peptide, particularly with regard to the ordering of the N-terminus compared to the disorder of the C-terminus. The temperature dependent VCD and IR spectra gave information about the melting behavior of different segments of the peptide. Spectral data analyses showed different transition temperatures for differently labeled peptides. Our study opened a new window for identifying segments in a secondary structure by promoting VCD into a true segment-specific conformational analysis tool.
Another study was carried out to identify the development of 310-helices, using model peptides containing high percentages of (-Me)Ala residues, which are known to promote 310-helices. Our results showed that in many cases it was not possible to distinguish 310- from - conformations by ECD data alone but that additional data from VCD is very useful for such interpretations. Further, we have shown that none of the factors as solvent, concentration, length, amphipathy can be considered as a unique factor for stabilizing a 310-helical conformation.