Homologous structural domains in chicken egg-white ovomucoid: Characterization and acid denaturation

The primary structure of ovomucoid shows considerable sequence homology at three contiguous regions which form structural domains I, II and III. In order to see whether or not the three domains fold similarly and acquire similar overall native conformation/shape, two fragments A and C were obtained by controlled peptic digestion of ovomucoid. The two fragments were investigated for their chemical composition, molecular weight, anti-tryptic activity, hydrodynamic behaviour, optical properties and acid denaturation. Results on molecular weight, amino acid composition and inhibitory acitivity show that the fragments A and C correspond respectively to domain I-II and domain III. Optical data suggested more exposure of tyrosine residues in the fragments than in the intact molecule. Domain III exists in a compact and globular conformation under native conditions whereas domain I-II and ovomucoid appear to possess asymmetric conformation. Results on acid denaturation show that the process is thermodynamically reversible and that inter-domain separation probably precedes denaturation of domains during acidification of ovomucoid.     

A developmentally regulated disappearance of slow myosin in fast-type muscles of the mouse

Histochemistry and immunocytochemistry using an antibody to adult rat slow-type myosin demonstrated that about 10% of the fibers in the mouse extensor digitorum longus and semimembranosus muscles contain slow myosin during the first month after birth. In adult animals, these muscles have only 0-08% slow myosin-containing fibers. These results demonstrate a developmentally linked disappearance of an adult-type myosin, and show that the adult phenotype of muscle fibers is not necessarily determined before birth as previously suggested.     

Secondary structural features of modules M2 and M3 of barnase in solution by NMR experiment and distance geometry calculation

Proteins consist of structural units such as globular domains, secondary structures, and modules. Modules were originally defined by partitioning a globular domain into compact regions, each of which is a contiguous polypeptide segment having a compact conformation. Since modules show close correlations with the intron positions of genes, they are regarded as primordial polypeptide pieces encoded by exons and shuffled, leading to yield new combination of them in early biological evolution. Do modules maintain their native conformations in solution when they are excised at their boundaries? In order to find answers to this question, we have synthesized modules of barnase, one of the bacterial RNases, and studied the solution structures of modules M2 (amino acid residues 24–52) and M3 (52–73) by 2D NMR studies. Some local secondary structures, α-helix, and β-turns in M2 and β-turns in M3, were observed in the modules at the similar positions to those in the intact barnase but the overall state seems to be in a mixture of random and native conformations. The present result shows that the excised modules have propensity to form similar secondary structures to those of the intact barnase. © 1993 Wiley-Liss, Inc.     

Conformational studies of Lewis X and Lewis A trisaccharides using NMR residual dipolar couplings.

The conformations of the histo-blood group carbohydrate antigens Lewis X (Le(x)) and Lewis A (Le(a)) were studied by NMR measurements of one-bond C-H residual dipolar couplings in partially oriented liquid crystal solutions. A strategy for rapid calculation of the difference between theoretical and experimental dipolar couplings of a large number of model structures generated by computer simulations was developed, resulting in an accurate model structure for the compounds. Monte Carlo simulations were used to generate models for the trisaccharides, and orientations of each model were sought that could reproduce the experimental residual dipolar coupling values. For both, Le(a) and Le(x), single low energy models giving excellent agreement with experiment were found, implying a compact rigidly folded conformation for both trisaccharides. The new approach was also applied to the pentasaccharides lacto-N-fucopentaose 2 (LNF-2) and lacto-N-fucopentaose 3 (LNF-3) proving its consistency and robustness. For describing the conformation of tightly folded oligosaccharides, a definition for characterization of ring planes in pyranoside chairs is proposed and applied to the analysis of the relation between the fucose and galactose residues in the epitopes, revealing the structural similarity between them.     

Phosphorylation alters backbone conformational preferences of serine and threonine peptides

Despite the notion that a control of protein function by phosphorylation works mainly by inducing its conformational changes, the phosphorylation effects on even small peptide conformation have not been fully understood yet. To study its possible effects on serine and threonine peptide conformations, we recently carried out pH- and temperature-dependent circular dichroism (CD) as well as (1)H NMR studies of the phosphorylated serine and threonine peptides and compared them with their unphosphorylated analogs. In the present article, by performing the self-consistent singular value decomposition analysis of the temperature-dependent CD spectra and by analyzing the (3)J(H(N),H(α)) coupling constants extracted from the NMR spectra, the populations of the polyproline II (PPII) and β-strand conformers of the phosphorylated Ser and Thr peptides are determined. As temperature is increased, the β-strand populations of both phosphorylated serine and threonine peptides increase. However, the dependences of PPII/β-strand population ratio on pH are different for these two cases. The phosphorylation of the serine peptide enhances the PPII propensity, whereas that of the threonine peptide has the opposite effect. This suggests that the serine and threonine phosphorylations can alter the backbone conformational propensity via direct but selective intramolecular hydrogen-bonding interactions with the peptide N--H groups. This clearly indicates that the phosphoryl group actively participates in modulating the peptide backbone conformations.     

Synthesis and C(2) -symmetric structure of a cyclotetrapeptide composed of anthranilic acid and leucine

A chiral cyclotetrapeptide (1) was synthesized from 2-nitrobenzoic acid and leucine. A single-crystal X-ray of the compound revealed a C(2) -symmetric bowl-shaped structure. The cyclic compound had a unique hydrogen-bonding network composed of three-centered hydrogen bonds and bifurcated hydrogen bonds between NH and CO of anthranilic residue. The NMR spectra and molecular modeling of 1 also suggested the chiral bowl structure.     

β‐amino acid substitution to investigate the recognition of angiotensin II (AngII) by angiotensin converting enzyme 2 (ACE2)

In spite of the important role of angiotensin converting enzyme 2 (ACE2) in the cardiovascular system, little is known about the substrate structural requirements of the AngII–ACE2 interaction. Here we investigate how changes in angiotensin II (AngII) structure affect binding and cleavage by ACE2. A series of C3 β‐amino acid AngII analogs were generated and their secondary structure, ACE2 inhibition, and proteolytic stability assessed by circular dichroism (CD), quenched fluorescence substrate (QFS) assay, and LC‐MS analysis, respectively. The β‐amino acid‐substituted AngII analogs showed differences in secondary structure, ACE2 binding and proteolytic stability. In particular, three different subsets of structure‐activity profiles were observed corresponding to substitutions in the N‐terminus, the central region and the C‐terminal region of AngII. The results show that β‐substitution can dramatically alter the structure of AngII and changes in structure correlated with ACE2 inhibition and/or substrate cleavage. β‐amino acid substitution in the N‐terminal region of AngII caused little change in structure or substrate cleavage, while substitution in the central region of AngII lead to increased β‐turn structure and enhanced substrate cleavage. β‐amino acid substitution in the C‐terminal region significantly diminished both secondary structure and proteolytic processing by ACE2. The β‐AngII analogs with enhanced or decreased proteolytic stability have potential application for therapeutic intervention in cardiovascular disease. Copyright © 2010 John Wiley & Sons, Ltd.     

Sexual dimorphism in Hucul horses using discriminant analysis

The purpose of this study has been to evaluate the applicability of discriminant function analysis to determine gender dimorphism in Hucul horses, based on morphological indices obtained in different stages of life. A total of 243 horses, divided into six age groups, have been examined in its course. For each horse we have measured 12 metric traits, which were then used to calculate 13 biometric indices commonly used in horse breeding in Poland. These have become the basis for defining functions classifying the animals by gender in each of the six age groups. This study answers the question of what parameters play the greatest role in the course of shaping of body proportions of male and female horses in post-foetal development. The following indices have been found to significantly contribute in discriminant models: boniness, smaller trunk length, height at the croup, pelvis width and width of chest.     

Ticks feeding on northern pocket gophers (Thomomys talpoides) in central Saskatchewan and the unexpected detection of Ixodes scapularis larvae

Morphological examination of ticks feeding on northern pocket gophers, Thomomys talpoides, near Clavet (Saskatchewan, Canada) revealed the presence of two genera, Ixodes and Dermacentor. All adult ticks collected were identified as I. kingi. Single strand conformation polymorphism (SSCP) analyses and DNA sequencing of the mitochondrial 16S rRNA gene confirmed the species identity of most Ixodes immatures as I. kingi (two nymphs and 82 larvae), and the Dermacentor immatures as D. variabilis (one nymph and one larva) and D. andersoni (three larvae). Six Ixodes larvae feeding on three T. talpoides individuals were identified as four different 16S haplotypes of I. scapularis, which was unexpected because there are no known established populations of this species in Saskatchewan. However, flagging for questing ticks and further examination of the ticks feeding on T. talpoides in two subsequent years failed to detect the presence of I. scapularis near Clavet, suggesting that there is no established population of I. scapularis in this area. Nonetheless, since I. scapularis is a vector of pathogenic agents, passive and active surveillance needs to be conducted in Saskatchewan on an ongoing basis to determine if this tick species and its associated pathogens become established within the province.