A possible novel function of dominance behaviour in queen-less colonies of the primitively eusocial wasp Ropalidia marginata

Unlike the queens of other primitively eusocial species, Ropalidia marginata queens are strikingly docile and non-aggressive individuals, never at the top of the behavioural dominance hierarchy of their colonies. Nevertheless, these queens are completely successful at suppressing worker reproduction, suggesting that they do not use aggression but employ some other mechanism (e.g. pheromones) to do so. Upon removal of the queen from a colony, a single worker, the 'potential queen', immediately begins to display highly elevated levels of aggression towards her nest mates. This individual becomes the next docile queen if the original queen is not returned. We attempt to understand the function of the temporary and amplified dominance behaviour displayed by the potential queen. We find that the dominance behaviour shown by the potential queen is unrelated to the number of her nest mates, their dominance ranks or ovarian condition. This suggests that aggression may not be used to actively suppress other workers and counter threat. Instead we find evidence that dominance behaviour is required for the potential queen's rapid ovarian development, facilitating her speedy establishment as the sole reproductive individual in the colony.     

Structural details on the binding of antihypertensive drugs captopril and enalaprilat to human testicular angiotensin I-converting enzyme

Angiotensin converting enzyme (ACE) plays a critical role in the circulating or endocrine renin-angiotensin system (RAS) as well as the local regulation that exists in tissues such as the myocardium and skeletal muscle. Here we report the high-resolution crystal structures of testis ACE (tACE) in complex with the first successfully designed ACE inhibitor captopril and enalaprilat, the Phe-Ala-Pro analogue. We have compared these structures with the recently reported structure of a tACE-lisinopril complex [Natesh et al. (2003) Nature 421, 551-554]. The analyses reveal that all three inhibitors make direct interactions with the catalytic Zn(2+) ion at the active site of the enzyme: the thiol group of captopril and the carboxylate group of enalaprilat and lisinopril. Subtle differences are also observed at other regions of the binding pocket. These are compared with N-domain models and discussed with reference to published biochemical data. The chloride coordination geometries of the three structures are discussed and compared with other ACE analogues. It is anticipated that the molecular details provided by these structures will be used to improve the binding and/or the design of new, more potent domain-specific inhibitors of ACE that could serve as new generation antihypertensive drugs.     

Roles of individual enzyme-substrate interactions by alpha-1,3-galactosyltransferase in catalysis and specificity

The retaining glycosyltransferase, alpha-1,3-galactosyltransferase (alpha3GT), is mutationally inactivated in humans, leading to the presence of circulating antibodies against its product, the alpha-Gal epitope. alpha3GT catalyzes galactose transfer from UDP-Gal to beta-linked galactosides, such as lactose, and in the absence of an acceptor substrate, to water at a lower rate. We have used site-directed mutagenesis to investigate the roles in catalysis and specificity of residues in alpha3GT that form H-bonds as well as other interactions with substrates. Mutation of the conserved Glu(317) to Gln weakens lactose binding and reduces the k(cat) for galactosyltransfer to lactose and water by 2400 and 120, respectively. The structure is not perturbed by this substitution, but the orientation of the bound lactose molecule is changed. The magnitude of these changes does not support a previous proposal that Glu(317) is the catalytic nucleophile in a double displacement mechanism and suggests it acts in acceptor substrate binding and in stabilizing a cationic transition state for cleavage of the bond between UDP and C1 of the galactose. Cleavage of this bond also linked to a conformational change in the C-terminal region of alpha3GT that is coupled with UDP binding. Mutagenesis indicates that His(280), which is projected to interact with the 2-OH of the galactose moiety of UDP-Gal, is a key residue in the stringent donor substrate specificity through its role in stabilizing the bound UDP-Gal in a suitable conformation for catalysis. Mutation of Gln(247), which forms multiple interactions with acceptor substrates, to Glu reduces the catalytic rate of galactose transfer to lactose but not to water. This mutation is predicted to perturb the orientation or environment of the bound acceptor substrate. The results highlight the importance of H-bonds between enzyme and substrates in this glycosyltransferase, in arranging substrates in appropriate conformations and orientation for efficient catalysis. These factors are manifested in increases in catalytic rate rather than substrate affinity.     

Fungal foliar diseases of Rauwolfia serpentina in wild,its seasonal occurrence,seed transmission and disease management

Rauwolfia serpentina is an important medicinal herb. Studies were conducted to determine causal organisms of foliar diseases in R. serpentina in Bhadra Wildlife Sanctuary during 2006–2009. The foliar disease incidence and its distribution and disease severity in nine state forest regions of the sanctuary were determined. The seedborne nature and transmission of the causal organism was also determined. The management of seedborne inoculum was done by seed dressing with fungicides. The effect of foliage infection on secondary metabolite content was also determined. Results of the present study indicated that Cercospora rauwolfiae is major leaf spot disease causing pathogen. The disease is homogeneously distributed through the study area. The foliar disease severity was high in Kagemanegiri forest during October–November. The minor leaf spot disease caused by Alternaria alternata occurred occasionally. Alternaria alternata is seedborne and seed transmitted and could be managed by seed treatment with Captra or Hyzeb. The secondary metabolites like alkaloids and steroids decreased with increase in foliar infection by C. rauwolfiae, while phenol and flavonoid contents increased. The study suggested that R. serpentina is affected in wild by C. rauwolfiae and A. alternata. The latter pathogen is seedborne and seed transmitted and controlled by seed treatment.     

CD45-mediated signaling pathway is involved in Rhizoctonia bataticola lectin (RBL)-induced proliferation and Th1/Th2 cytokine secretion in human PBMC

We earlier reported the mitogenic and immunostimulatory activities of Rhizoctonia bataticola lectin (RBL), purified from phytopathogenic fungus R. bataticola in human PBMC. The lectin demonstrates specificity towards glycoproteins containing complex N-glycans. Since CD45-protein tyrosine phosphatase that abundantly expresses N-glycans is important in T-cell signaling, the study aimed to investigate the involvement of CD45 in the immunomodulatory activities of RBL. Flowcytometry and confocal microscopy studies revealed that RBL exhibited binding to PBMC and colocalized with CD45. The binding was comparable in cells expressing different CD45 isoforms-RA, -RB and -RO. CD45 blocking antibody reduced the binding and proliferation of PBMC induced by RBL. CD45-PTPase inhibitor dephostatin inhibited RBL-induced proliferation, expression of CD25 and pZAP-70. RBL-induced secretion of Th1/Th2 cytokines were significantly inhibited in presence of dephostatin. Also, dephostatin blocked phosphorylation of p38MAPK and STAT-5 that was crucial for the biological functions of RBL. The study demonstrates the involvement of CD45-mediated signaling in RBL-induced PBMC proliferation and Th1/Th2 cytokine secretion through activation of p38MAPK and STAT-5.     

Basic and applied research in India: Present and future

Models for supporting organized applied research programs Position of India in the global scenario of basic research New policy initiatives and programs Conclusion     

Molecular-genetic analysis of two cases with retinoblastoma: Benefits for disease management

Effective counselling and management of retinoblastoma families using genetic information is presently practised in many parts of the world. We studied histopathological, chromosomal and molecular-genetic data of two retinoblastoma patients from India. The two patients, one with bilateral and the other with unilateral retinoblastoma, underwent complete ophthalmic examination, cytogenetic study, retinoblastoma gene (RB1) mutational analysis andRB1 promoter region methylation screening. In the bilateral retinoblastoma patient deletion of chromosome region 13q14 in peripheral blood lymphocytes and a hemizygous novel 8-bp deletion in exon 4 ofRB1 in tumour sample were observed. In the unilaterally affected patient CGA to TGA transition protein truncation mutations were observed in exons 8 and 14 ofRB1.     

Structure of UDP complex of UDP-galactose:beta-galactoside-alpha -1,3-galactosyltransferase at 1.53-A resolution reveals a conformational change in the catalytically important C terminus

UDP-galactose:beta-galactosyl alpha-1,3-galactosyltransferase (alpha3GT) catalyzes the transfer of galactose from UDP-alpha-d-galactose into an alpha-1,3 linkage with beta-galactosyl groups in glycoconjugates. The enzyme is expressed in many mammalian species but is absent from humans, apes, and old world monkeys as a result of the mutational inactivation of the gene; in humans, a large fraction of natural antibodies are directed against its product, the alpha-galactose epitope. alpha3GT is a member of a family of metal-dependent retaining glycosyltransferases including the histo-blood group A and B synthases. A crystal structure of the catalytic domain of alpha3GT was recently reported (Gastinel, L. N., Bignon, C., Misra, A. K., Hindsgaul, O., Shaper, J. H., and Joziasse, D. H. (2001) EMBO J. 20, 638-649). However, because of the limited resolution (2.3 A) and high mobility of the atoms (as indicated by high B-factors) this structure (form I) does not provide a clear depiction of the catalytic site of the enzyme. Here we report a new, highly ordered structure for the catalytic domain of alpha3GT at 1.53-A resolution (form II). This provides a more accurate picture of the details of the catalytic site that includes a bound UDP molecule and a Mn(2+) cofactor. Significantly, in the new structure, the C-terminal segment (residues 358-368) adopts a very different, highly structured conformation and appears to form part of the active site. The properties of an Arg-365 to Lys mutant indicate that this region is important for catalysis, possibly reflecting its role in a donor substrate-induced conformational change.     

Roles of active site tryptophans in substrate binding and catalysis by alpha-1,3 galactosyltransferase

Aromatic amino acids are frequent components of the carbohydrate binding sites of lectins and enzymes. Previous structural studies have shown that in alpha-1,3 galactosyltransferase, the binding site for disaccharide acceptor substrates is encircled by four tryptophans, residues 249, 250, 314, and 356. To investigate their roles in enzyme specificity and catalysis, we expressed and characterized variants of the catalytic domain of alpha-1,3 galactosyltransferase with substitutions for each tryptophan. Substitution of glycine for tryptophan 249, whose indole ring interacts with the nonpolar B face of glucose or GlcNAc, greatly increases the K(m) for the acceptor substrate. In contrast, the substitution of tyrosine for tryptophan 314, which interacts with the beta-galactosyl moiety of the acceptor and UDP-galactose, decreases k(cat) for the galactosyltransferase reaction but does not affect the low UDP-galactose hydrolase activity. Thus, this highly conserved residue stabilizes the transition state for the galactose transfer to disaccharide but not to water. High-resolution crystallographic structures of the Trp(249)Gly mutant and the Trp(314)Tyr mutant indicate that the mutations do not affect the overall structure of the enzyme or its interactions with ligands. Substitutions for tryptophan 250 have only small effects on catalytic activity, but mutation of tryptophan 356 to threonine reduces catalytic activity for both transferase and hydrolase activities and reduces affinity for the acceptor substrate. This residue is adjacent to the flexible C-terminus that becomes ordered on binding UDP to assemble the acceptor binding site and influence catalysis. The results highlight the diverse roles of these tryptophans in enzyme action and the importance of k(cat) changes in modulating glycosyltransferase specificity.