Changes to near-surface region of PTFE during dry sliding against steel

PTFE specimens were slid against an EN24 disc. The unworn and worn surfaces as well as the wear debris were examined by X-ray diffraction. Sliding was found to introduce (i) shrinkage of the unit cell, (ii) enlargement of crystallites and (iii) residual stresses in the slid PTFE surface. No conformational changes in the 157 helix could be observed due to sliding. The wear debris was found to be 1 mgrm thick warped laminates.

Crystal and molecular structure of the ammonium salt of the dinucleoside monophosphate d(CpG)

The crystal and molecular structure of the ammonium salt of deoxycytidylyl-(3'-5')-deoxyguanosine has been determined from 0.85 A resolution single crystal X-ray diffraction data. The crystals obtained by acetone diffusion technique at -20 degrees C, are orthorhombic, P212121, a = 12.880(2), b = 17444(2) and c = 27.642(2) A. The structure was solved by high resolution Patterson and Fourier methods and refined to R = 0.136. There are two d(CpG) molecules in the asymmetric unit forming a mini left handed Z-DNA helix. This is in contrast to the earlier reported forms of d(CpG) where the molecules form self base paired duplexes. There are two ammonium ions in the asymmetric unit. The major groove NH+4 ion interacts with N7 of guanines through water bridges besides making H-bonded interactions directly with the phosphate oxygen atoms. A second NH+4 ion is found in the minor groove interacting directly with the phosphate oxygen atoms. Symmetry related molecules pack in such a way that the cytosine base stacks on cytosine and guanine base on guanine. Our structure demonstrates that alternating d(CpG) sequences have the ability to adopt the left handed Z-DNA structure even at the dimer level i.e., in a sequence which is only two base pairs long.

Some implications of an alternative structure for DNA

We have constructed a space-filling (Corey-Pauling-Koltun) model of an alternative structure for DNA. This structure is not a double helix, but consists of a pair of polynucleotide strands lying side by side and held together by Watson-Crick base pairing. Each of the two strands has alternating right- and left-handed helical segments approximately five base pairs in length. Sugar residues in alternating segments along a strand point in opposite directions. A structure slightly different from the present one proposed earlier by ourselves and another group and in which sugars in a strand all point in the same direction is ruled out. The present structure yields natural solutions to the problems of supercoiling of DNA and of strand separation during DNA replication. This model is energetically more favorable than the double helix.

Stereochemistry of collagen

This review article, based on a lecture delivered in Madras in 1985, is an account of the author's experience in the working out of the molecular structure and conformation of the collagen triple-helix over the years 1952–78. It starts with the first proposal of the correct triple-helix in 1954, but with three residues per turn, which was later refined in 1955 into a coiled-coil structure with approximately 3.3 residues per turn. The structure readily fitted proline and hydroxyproline residues and required glycine as every third residue in each of the three chains. The controversy regarding the number of hydrogen bonds per tripeptide could not be resolved by X-ray diffraction or energy minimization, but physicochemical data, obtained in other laboratories during 1961–65, strongly pointed to two hydrogen bonds, as suggested by the author. However, it was felt that the structure with one straight NH … O bond was better. A reconciliation of the two was obtained in Chicago in 1968, by showing that the second hydrogen bond is via a water molecule, which makes it weaker, as found in the physicochemical studies mentioned above. This water molecule was also shown, in 1973, to take part in further cross-linking hydrogen bonds with the OH group of hydroxyproline, which occurred always in the location previous to glycine, and is at the right distance from the water. Thus, almost all features of the primary structure, X-ray pattern, optical and hydrodynamic data, and the role of hydroxyproline in stabilising the triple helical structure, have been satisfactorily accounted for. These also lead to a confirmation of Pauling's theory that vitamin C improves immunity to diseases, as explained in the last section.

Interaction of metal ions with nucleic acids and related compounds. II. Studies on Au(III)-nucleic acid system

The nature of interaction of Au(III) with nucleic acids was studied by using methods such as uv and ir spectrophotometry, viscometry, pH titrations, and melting-temperature measurements. Au(III) is found to interact slowly with nucleic acids over a period of several hours. The uv spectra of native calf-thymus DNA 9pH 5.6 acetate buffer containing (0.01M NaCIO4) showed a shift in λ max to high wavelengths and an increase in optical density at 260 nm. There was a fourfold decrease in viscosity (expressed as ηsp/c). The reaction was faster at pH 4.0 and also with denatured DNA (pH 5.6) and whole yeast RNA (pH 5.6). The order of preference of Au(III) (as deduced from the time of completion of reaction) for the nucleic acids in RNA > denatured DNA > DNA. The reaction was found to be completely reversible with respect KCN. Infrared spectra of DNA-Au(III) complexes showed binding to both the phosphate and bases of DNA. The same conclusions were also arrived at by melting-temperature studies of Au(III)-DNA system. pH titrations showed liberation of two hydroxylions at r = 0.12 [r = moles of HAuCl4 added per mole of DNA-(P)] and one hydrogen ion at r = 0.5. The probable binding sites could be N(1)/N(7) of adenine, N(7) and/or C(6)O of guanine, N(3) of cytosine and N(3) of thymine. DNAs differing in their (G = C)-contents [Clostridium perfingens DNA(G = C, 29%), salmon sperm DNA (G + C, 42%) and Micrococcus lysodeikticus DNA(G + C, 29%), salmon sperm DNA (G = C, 72%)] behaved differently toward Au(III). The hyperchromicity observed for DNAs differing in (G + C)-content and cyanide reversal titrations indicate selectivity toward ( A + T)-rich DNA at lw values of r. Chemical analysis and job's continuous variation studies indicated the existence of possible complexes above and below r = 1. The results indicate that Au(III) ions probably bind to hte phosphate group in the initial stages of the reaction, particularly at low values of r, and participation of the base interaction also increases. Cross-linking of the two strands by Au(III) may take place, but a complete collapse of the doulbe helix is not envisaged. It is probable that tilting of the bases or rotaiton of the bases around the glucosidic bond, resulting in a significant distrotion of the double helix, might take place due to binding of Au(III) to DNA.

A helical peptide containing a majority of valyl residues. The crystal structure of t-butyloxy- carbonyl-(L-valyl-_-aminoisobutyryl)3-L-valyl methyl ester onohydrate

The monohydrate of the heptapeptide t-butyloxycarbonyl-(L-valyl-α-aminoiso-butyryl)3-L-valyl methyl ester crystallizes in the orthorhombic space group P212121 with four molecules in a unit cell with the dimensions α= 9.375, b = 19.413 and c = 25.878 ÅA. The structure has been solved by direct methods and refined to an R value of 0.059 for 3633 observed reflections. The molecule in the structure exists as a slightly distorted 310-helix stabilized by five 4 -> 1 intramolecular hydrogen bonds, indicating the overwhelming influence of α-aminoisobutyryl (Aib) residues in dictating helical fold even when a majority of residues in the peptide have a low intrinsic propensity to be in helices. Contrary to what is expected in helical structures, the valyl side chains, two of which are disordered, exhibit all three possible conformations. The molecules arrange themselves in a head-to-tail fashion along the c-axis. The columns thus generated pack nearly hexagonally in the crystal.

Empirical torsional potential functions from protein structure data. Phi- and psi-potentials for non-glycyl amino acid residues.

The torsional potential functions Vt(phi) and Vt(psi) around single bonds N--C alpha and C alpha--C, which can be used in conformational studies of oligopeptides, polypeptides and proteins, have been derived, using crystal structure data of 22 globular proteins, fitting the observed distribution in the (phi, psi)-plane with the value of Vtot(phi, psi), using the Boltzmann distribution. The averaged torsional potential functions, obtained from various amino acid residues in L-configuration, are Vt(phi) = 1.0 cos (phi + 60 degrees); Vt(psi) = 0.5 cos (psi + 60 degrees) - 1.0 cos (2 psi + 30 degrees) - 0.5 cos (3 psi + 30 degrees). The dipeptide energy maps Vtot(phi, psi) obtained using these functions, instead of the normally accepted torsional functions, were found to explain various observations, such as the absence of the left-handed alpha helix and the C7 conformation, and the relatively high density of points near the line psi = 0 degrees. These functions derived from observational data on protein structures, will, it is hoped, explain various previously unexplained facts in polypeptide conformation.

A nonhelical, multiple β-turn conformation in a glycine-rich heptapeptide fragment of trichogin A IV containing a single central α-aminoisobutyric acid residue

The conformational properties of the protected seven-residue C-terminal fragment the lipopeptaibol antibiotic Trichogin A IV (Boc-Gly-Gly-Leu-Aib-Gly-Ile-Leu-OMe) has been examined in CDCl3 and (CD3)2SO by 1H-nmr. Evidence for a multiple β-turn conformation [type I′ at Gly(1)-Gly(2), type II at Leu(3)-Aib(4), and a type I′ at Aib(4)-Gly(5)] suggests that Leu(3) has preferred an extended or semiextended conformation over a helical conformation in CDCl3. This structure is thus in contrast to earlier observations of seven-residue peptides containing a single central Aib preferring helical conformations in both solution and crystalline slates. A structural transition to a frayed right-handed helix is absented in (CD3)2SO. These results suggest that nonhelical conformations may be important in Gly-rich peptides containing Aib. Further, the presence of amino acids with contradictory influences on backbone conformational freedom can lead to well-defined conformational transitions even in small peptides

A C-H…O hydrogen bond stabilized polypeptide chain reversal motif at the C-terminus helices in proteins.

The serendipitous observation of a C–Hcdots, three dots, centeredO hydrogen bond mediated polypeptide chain reversal in synthetic peptide helices has led to a search for the occurrence of a similar motif in protein structures. From a dataset of 634 proteins, 1304 helices terminating in a Schellman motif have been examined. The C–Hcdots, three dots, centeredO interaction between the T−4 CαH and T+1 C=O group (Ccdots, three dots, centeredO≤3.5 Å) becomes possible only when the T+1 residue adopts an extended β conformation (T is defined as the helix terminating residue adopting an αL conformation). In all, 111 examples of this chain reversal motif have been identified and the compositional and conformational preferences at positions T−4, T, and T+1 determined. A marked preference for residues like Ser, Glu and Gln is observed at T−4 position with the motif being further stabilized by the formation of a side-chain–backbone Ocdots, three dots, centeredH–N hydrogen bond involving the side-chain of residue T−4 and the N–H group of residue T+3. In as many as 57 examples, the segment following the helix was extended with three to four successive residues in β conformation. In a majority of these cases, the succeeding β strand lies approximately antiparallel with the helix, suggesting that the backbone C–Hcdots, three dots, centeredO interactions may provide a means of registering helices and strands in an antiparallel orientation. Two examples were identified in which extended registry was detected with two sets of C–Hcdots, three dots, centeredO hydrogen bonds between (T−4) CαHcdots, three dots, centeredC=O (T+1) and (T−8) CαHcdots, three dots, centeredC=O (T+3).

Probing the role of the C-H center dot center dot center dot O hydrogen bond stabilized polypeptide chain reversal at the C-terminus of designed peptide helices. Structural characterization of three decapeptides

The structural characterization in crystals of three designed decapeptides containing a double D-segment at the C-terminus is described. The crystal structures of the peptides Boc-Leu-Aib-Val-Xxx-Leu-Aib-Val- (D)Ala-(D)Leu-Aib-OMe, (Xxx = Gly 2, (D)Ala 3, Aib 4) have been determined and compared with those reported earlier for peptide 1 (Xxx = Ala) and the all L analogue Boc-Leu-Aib-Val-Ala-Leu-Aib-Val-Ala-Leu-Aib-OMe, which yielded a perfect right-handed a-helical structure. Peptides 1 and 2 reveal a right-handed helical segment spanning residues 1 to 7, ending in a Schellman motif with Ala(8) functioning as the terminating residue. Polypeptide chain reversal occurs at residue 9, a novel feature that appears to be the consequence of a C-(HO)-O-... hydrogen bond between residue 4 (CH)-H-alpha and residue 9 CO groups. The structures of peptides 3 and 4, which lack the pro R hydrogen at the C-alpha atom of residue 4, are dramatically different. Peptide 3 adopts a right-handed helical conformation over the 1 to 7 segment. Residues 8 and 9 adopt at conformations forming a C-terminus type I' beta-turn, corresponding to an incipient left-handed twist of the polypeptide chain. In peptide 4, helix termination occurs at Aib(6), with residues 6 to 9 forming a left-handed helix, resulting in a structure that accommodates direct fusion of two helical segments of opposite twist. Peptides 3 and 4 provide examples of chiral residues occurring in the less favored sense of helical twist; (D)Ala(4) in peptide 3 adopts an alpha(R) conformation, while (L)Val(7) in 4 adopts an alpha(L) conformation. The structural comparison of the decapeptides reported here provides evidence for the role of specific C-(HO)-O-... hydrogen bonds in stabilizing chain reversals at helix termini, which may be relevant in aligning contiguous helical and strand segments in polypeptide structures.

Diproline Templates as Folding Nuclei in Designed Peptides. Conformational Analysis of Synthetic Peptide Helices Containing Amino Terminal Pro-Pro Segments

The effect of N-terminal diproline segments in nucleating helical folding in designed peptides has been studied in two model sequences Piv-Pro-Pro-Aib-Leu-Aib-Phe-OMe (1) and Boc-Aib-Pro-Pro-Aib-Val-Ala-Phe-OMe (2). The structure of 1 in crystals, determined by X-ray diffraction, reveals a helical (RR) conformation for the segment residues 2 to 5, stabilized by one 4 -> 1 hydrogen bond and two 5 -> 1 interactions. The N-terminus residue, Pro(1) adopts a polyproline II (P-II) conformation. NMR studies in three different solvent systems support a conformation similar to that observed in crystals. In the apolar solvent CDCl3, NOE data favor the population of both completely helical and partially unfolded structures. In the former, the Pro-Pro segment adopts an alpha(R)-alpha(R) conformation, whereas in the latter, a P-II-alpha(R) structure is established. The conformational equilibrium shifts in favor of the P-II-alpha(R) structure in solvents like methanol and DMSO. A significant population of the Pro(1)- Pro(2) cis conformer is also observed. The NMR results are consistent with the population of at least three conformational states about Pro- Pro segment: trans alpha(R)-alpha(R), trans P-II-alpha(R) and cis P-II-alpha(R). Of these, the two trans conformers are in rapid dynamic exchange on the NMR time scale, whereas the interconversion between cis and trans form is slow. Similar results are obtained with peptide 2. Analysis of 462 diproline segments in protein crystal structures reveals 25 examples of the alpha(R)-alpha(R) conformation followed by a helix. Modeling and energy minimization studies suggest that both P-II-alpha(R) and alpha(R)-alpha(R) conformations have very similar energies in the model hexapeptide 1

Structural Investigations on Poly(4-hydroxy-L-proline). 2. Physicochemical Studies

We report experimental studies which confirm our prediction, namely that the ordered structure of poly(hydroxypro1ine) in solution corresponds to a left-handed helical structure with intrachain hydrogen bonds. The CD studies show that the poly(hydroxypro1ine) molecule has essentially the same conformation in aqueous solution and in the film obtained subsequently by evaporation. X-ray diffraction patterns of the sample in this form (B form) have been recorded at different relative humidities. The patterns recorded at relative humidities over 66% can be interpreted in terms of a helical structure with intrachain hydrogen bonds. These results lead us to conclude that the ordered conformation of poly(hydroxypro1ine) in solution is form B and not form A. This offers a simple explanation for the greater stability of the poly(hydroxypro1ine) helix in solution as compared to the poly(pro1ine) form I1 helix and also for the absence of mutarotation for poly(hydroxypro1ine).

Kinetic studies on the interaction of gold(III) with nucleic acids. I. Native DNA-Au(III) system-spectrophotometric studies

Kinetics of the interaction of Au(III) with native calf thymus DNA has been studied spectrophotometrically to determine the kinetic parameters and to examine their dependency on the concentrations of DNA and Au(III), temperature, ionic strength and pH. The reaction is of the first order with respect to both the nucleotide unit of DNA and Au(III) in the stoichiometry of 2∶1 respectively. The rate constants vary with the initial ratio of DNA to Au(III) and is attributed to the effect of free chloride ions and the existence of a number of reaction sites with slight difference in the rate constants. The activation energies of this interaction have been found to be 14–16 kcal/mol. From the effect of ionic strength the reaction is found to occur between a positive and a negative ion in the rate-limiting step. The logarithm of rate constants are the linear function of pH and the slopes are dependent on ther-values. A plausible mechanism has been proposed which involves a primary dissociation of the major existing species (AuCl2(OH)2)−, to give (AuCl2)+ which then reacts with a site in the nucleotide unit of DNA in the rate-liminting step followed by a rapid binding to another site on the complementary strand of the DNA double helix. There exist a number of binding sites with slight difference in reactivity.

Fourfold Helical Structures for Polypeptides

Fourfold helical structures for polypeptides and their association in regular lattices with interchain hydrogen bonds were investigated by model building studies. These studies revealed that stereochemically satisfactory fourfold helical sturctures are possible for polyglycine, polyproline, and copolymers of glycine and proline with two and four units in the monomer. In these structures the unit height h for the backbone has been found to be restricted from 2.7 to 3.1 k, with four peptide units per turn of the helix. Energetically both fourfold and threefold helical structures are equally favorable.

Empirical torsional potential functions from protiesn structure DATA Phi- and Psi-Potentials for Non-glycyl Amino Acid Residues

The torsional potential functions Vt(φ) and Vt(ψ) around single bonds N–Cα and Cα-C, which can be used in conformational studies of oligopeptides, polypeptides and proteins, have been derived, using crystal structure data of 22 globular proteins, fitting the observed distribution in the (φ, ψ)-plane with the value of Vtot(φ, ψ), using the Boltzmann distribution. The averaged torsional potential functions, obtained from various amino acid residues in l-configuration, are Vt(φ) = – 1.0 cos (φ + 60°); Vt(ψ) = – 0.5 cos (ψ + 60°) – 1.0 cos (2ψ + 30°) – 0.5 cos (3ψ + 30°). The dipeptide energy maps Vtot(φ, ψ) obtained using these functions, instead of the normally accepted torsional functions, were found to explain various observations, such as the absence of the left-handed alpha helix and the C7 conformation, and the relatively high density of points near the line ψ = 0°. These functions, derived from observational data on protein structures, will, it is hoped, explain various previously unexplained facts in polypeptide conformation.