C-H…O hydrogen bond mediated chain reversal in a peptide containing g-amino acid residue, determined directly from powder X-ray diffraction data

The finding that peptides containing -amino acid residues give rise to folding patterns hitherto unobserved in -amino acid peptides[1] has stimulated considerable interest in the conformational properties of peptides built from , and residues,[2] as the introduction of additional methylene (CH2) units into peptide chains provides further degrees of conformational freedom.

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.

Interactions of methoxyamine with pyridoxal-5'-phosphate-Schiff's base at the active site of sheep liver serine hydroxymethyltransferase

The mechanism of interaction of methoxyamine with sheep liver serine hydroxymethyltransferase (EC 2.1.2.1) (SHMT) was established by measuring changes in enzyme activity, visible absorption spectra, circular dichroism and fluorescence, and by evaluating the rate constant by stopped-flow spectrophotometry. Methoxyamine can be considered as the smallest substituted aminooxy derivative of hydroxylamine. It was a reversible noncompetitive inhibitor (Ki = 25 microM) of SHMT similar to O-amino-D-serine. Like in the interaction of O-amino-D-serine and aminooxyacetic acid, the first step in the reaction was very fast. This was evident by the rapid disappearance of the enzyme-Schiff base absorbance at 425 nm with a rate constant of 1.3 x 10(3) M-1 sec-1 and CD intensity at 430 nm. Concomitantly, there was an increase in absorbance at 388 nm (intermediate I). The next step in the reaction was the unimolecular conversion (1.1 x 10(-3) sec-1) of this intermediate to the final oxime absorbing at 325 nm. The identity of the oxime was established by its characteristic fluorescence emission at 460 nm when excited at 360 nm and by high performance liquid chromatography. These results highlight the specificity in interactions of aminooxy compounds with sheep liver serine hydroxymethyltransferase and that the carboxyl group of the inhibitors enhances the rate of the initial interaction with the enzyme.

Modes of binding of guanosine monophosphates to ribonuclease T1 - A computer modelling study

Computer-modelling studies on the modes of binding of the three guanosine monophosphate inhibitors 2'-GMP, 3'-GMP, and 5'-GMP to ribonuclease (RNase) T1 have been carried out by energy minimization in Cartesian-coordinate space. The inhibitory power was found to decrease in the order 2'-GMP > 3'-GMP > 5'-GMP in agreement with the experimental observations. The ribose moiety was found to form hydrogen bonds with the protein in all the enzyme-inhibitor complexes, indicating that it contributes to the binding energy and does not merely act as a spacer between the base and the phosphate moieties as suggested earlier. 2'-GMP and 5'-GMP bind to RNase T1 in either of the two ribose puckered forms (with C3'-endo more favoured over the C2'-endo) and 3'-GMP binds to RNase T1 predominantly in C3'-endo form. The catalytically important residue His-92 was found to form hydrogen bond with the phosphate moiety in all the enzyme-inhibitor complexes, indicating that this residue may serve as a general acid group during catalysis. Such an interaction was not found in either X-ray or two-dimensional NMR studies.

Structure of the putative mutarotase YeaD from Salmonella typhimurium in two different forms: in vivo binding of a sugar phosphate

Salmonella typhimurium YeaD (stYeaD), annotated as a putative aldose 1-epimerase, has a very low sequence identity to other well characterized mutarotases. Sequence analysis suggested that the catalytic residues and a few of the substrate-binding residues of galactose mutarotases (GalMs) are conserved in stYeaD. Determination of the crystal structure of stYeaD in an orthorhombic form at 1.9 angstrom resolution and in a monoclinic form at 2.5 angstrom resolution revealed this protein to adopt the beta-sandwich fold similar to GalMs. Structural comparison of stYeaD with GalMs has permitted the identification of residues involved in catalysis and substrate binding. In spite of the similar fold and conservation of catalytic residues, minor but significant differences were observed in the substrate- binding pocket. These analyses pointed out the possible role of Arg74 and Arg99, found only in YeaD-like proteins, in ligand anchoring and suggested that the specificity of stYeaD may be distinct from those of GalMs

New flame retardant polyarylazo phosphate and phosphoramide esters

Two series of flame retardant polymers, viz. polyarylazo phosphate and phosphoramide esters, were synthesized by solution polycondensation of 4,4′-dihydroxyazobenzene with various aryl phosphorodichlorides and aryl phosphoramidic dichlorides. They were characterized by i.r. 1H-, 13C- and 31P-NMR spectroscopy. The molar mass, thermal and flammability studies were carried out by viscometry, thermogravimetry and limiting oxygen index respectively to examine the influence of the phosphate and phosphoramide linkages. The polyphosphoramide esters possess better thermal and flammability characteristics than the polyphosphate esters.

Reactivity with hydrogen of pure iron oxide and of iron oxides doped with oxides of Mn, Co, Ni and Cu

Using dynamic TG in H2, X-ray powder diffraction and Mössbauer Spectroscopy the reactivities fot hydrogen reduction of Fe2O3 prepared at different temperatures, Fe2O3 doped with oxides of Mn, Co, Ni and Cu prepared at 300DaggerC from nitrate precursors and intermediate spinels derived from above samples during reduction have been explored. The reactivity is higher for finely divided Fe2O3 prepared at 250DaggerC. The reduction is retarded by Mn, marginally affected by Co and accelerated by Ni and Cu, especially at higher (5 at.%) dopant concentration. These reactivities confirmed also by isothermal experiments, are ascribed to the nature of disorder in the metastable intermediate spinels and to hydrogen rsquospill overrsquo effects.

Serine Hydroxymethyltransferase from Mung Bean (Vigna radiata) Is Not a Pyridoxal-5'-Phosphate- Dependent Enzyme

Serine hydroxymethyltransferase from mammalian and bacterial sources is a pyridoxal-5'-phosphate-containing enzyme, but the requirement of pyridoxal-5'-phosphate for the activity of the enzyme from plant sources is not clear. The specific activity of serine hydroxymethyltransferase isolated from mung bean (Vigna radiata) seedlings in the presence and absence of pyridoxal-5'-phosphate was comparable at every step of the purification procedure. The mung bean enzyme did not show the characteristic visible absorbance spectrum of pyridoxal-5'-phosphate protein. Unlike the enzymes from sheep, monkey, and human liver, which were converted to the apoenzyme upon treatment with L-cysteine and dialysis, the mung bean enzyme similarly treated was fully active. Additional evidence in support of the suggestion that pyridoxal-5'-phosphate may not be required for the mung bean enzyme was the observation that pencillamine, a well-known inhibitor of pyridoxal-5'-phosphate enzymes, did not perturb the enzyme spectrum or inhibit the activity of mung bean serine hydroxymethyltransferase. The sheep liver enzyme upon interaction with O-amino-D-serine gave a fluorescence spectrum with an emission maximum at 455 nm when excited at 360 nm. A 100-fold higher concentration of mung bean enzyme-O-amino-D-serine complex did not yield a fluorescence spectrum. The following observations suggest that pyridoxal-5'-phosphate normally present as a coenzyme in serine hydroxymethyltransferase was probably replaced in mung bean serine hydroxymethyltransferase by a covalently bound carbonyl group: (a) inhibiton by phenylhydrazine and hydroxylamine, which could not be reversed by dialysis and or addition of pyridoxal-5'-phosphate; (b) irreversible inactivation by sodium borohydride; (c) a spectrum characteristic of a phenylhydrazone upon interaction with phenylhydrazine; and (d) the covalent labeling of the enzyme with substrate/product serine and glycine upon reduction with sodium borohydride. These results indicate that in mung bean serine hydroxymethyltransferase, a covalently bound carbonyl group has probably replaced the pyridoxal-5'-phosphate that is present in the mammalian and bacterial enzymes.

Synthesis and spectral studies of polyamide-phosphate esters from phosphine-oxide-containing diols with aryl phosphorodichloridates and their thermal and flammability behaviour

Polyamide-phosphate esters were synthesized by interfacial polycondensation of aryl phosphorodichloridates with the diols of phenoxaphosphine and phosphine oxide in the presence of a phase-transfer catalyst. The polymers were characterized by infra-red and 1H, 13C and 31P nuclear magnetic resonance (n.m.r.) spectroscopy. The molecular weights were determined by end-group analysis using 31P n.m.r. spectral data. The phenoxaphosphine-containing polymers showed superior thermostability and flame retardancy over the phosphine-oxide-containing polymers.

Oxidation of aromatic substrates with hydrogen peroxide and hydrochloric acid

Chloroquinones are prepared conveniently from phenol, naphthols and aromatic amines.