Constitutively active mutant gives novel insights into the mechanism of bitter taste receptor activation

The human bitter taste receptors (T2Rs) belong to the G-protein coupled receptor (GPCR) superfamily. T2Rs share little homology with the large subfamily of Class A G-protein coupled receptors, and their mechanisms of activation are poorly understood. Guided by biochemical and molecular approaches, we identified two conserved amino acids Gly281·?? and Ser285?·?? present on transmembrane (TM) helices, TM1 and TM7, which might play important roles in T2R activation. Previously, it was shown that naturally occurring Gly511·?? mutations in the dim light receptor, rhodopsin, cause autosomal dominant retinitis pigmentosa, with the mutants severely defective in signal transduction. We mutated Gly281·?? and Ser285?·?? in T2R4 to G28A, G28L, S285A, S285T, and S285P, and carried out pharmacological characterization of the mutants. No major changes in signaling were observed upon mutation of Gly281·?? in T2R4. Interestingly, S285A mutant displayed agonist-independent activity (approximately threefold over basal wild-type T2R4 or S285T or S285P). We propose that Ser285?·?? stabilizes the inactive state of T2R4 by a network of hydrogen-bonds connecting important residues on TM1-TM2-TM7. We compare and contrast this hydrogen-bond network with that present in rhodopsin. Thus far, S285A is the first constitutively active T2R mutant reported, and gives novel insights into T2R activation.     

Characterization of silver nanoparticles synthesized by using marine isolate Streptomyces albidoflavus

Silver nanoparticles production by the green chemistry approach was investigated using an isolated marine actinomycetes strain. The isolated strain was identified as Streptomyces albidoflavus based on chemotaxonomic and ribotyping properties. The strain revealed production of silver nanoparticles both extracellular and intracellularly. Surface Plasmon Resonance analysis with the function of time revealed that particle synthesis by this strain is reaction time dependent. The produced particles were spherical shaped and monodispersive in nature and showed a single surface plasmon resonance peak at 410 nm. Size distribution histograms indicated production of 10-40- nm-size nanoparticles with a mean size of 14.5 nm. FT-IR spectra of nanopartilces showed N-H, C-H, and C-N stretching vibrations, denoting the presence of amino acid/ peptide compounds on the surface of silver nanoparticles produced by S. albidoflavus. Synthesized nanoparticles revealed a mean negative zeta potential and electrophoretic mobility of -8.5 mV and -0.000066 cm2/Vs, respectively. The nanoparticles produced were proteinaceous compounds as capping agents with -8.5 mV zeta potential and revealed antimicrobial activity against both Gram-negative and -positive bacterial strains. Owing to their small size, these particles have greater impact on industrial application spectra.     

Association of 1347 G/A cytochrome P450 4F2 (CYP4F2) gene variant with hypertension and stroke

Genetic variants of cytochrome P450 4F2 (CYP4F2) gene have been suggested to be risk factors for hypertension, cardiovascular diseases and stroke. In the present case–control study we investigated the association of 1347 G/A polymorphism (rs2108622) in the 11th exon region of CYP4F2 gene with hypertension, ischemic stroke and stroke subtypes classified according to TOAST (Trial of Org 10172 in Acute Stroke Treatment) classification. Five hundred and seven stroke patients (hypertensives: normotensives = 279:228) and four hundred and eighty seven, age and sex matched controls (males: females = 356:131) (hypertensives: normotensives = 148:339) were involved in the study. The genotypes were determined by PCR-RFLP technique. Genotypes were confirmed by subjecting the PCR products to sequencing. Significant difference was observed in the genotypic distribution and allelic frequency between the stroke patients and healthy controls. AA genotype and A allele associated significantly with stroke and hypertension [P = 0.009; OR = 1.59 (95% CI = 1.119–2.283) and P = 0.010; OR = 1.26 (95% CI = 1.056–1.502); P = 0.01; OR = 1.58 (95% CI = 1.11–2.272) and P = 0.010; OR = 1.25(95% CI = 1.054–1.504) respectively]. A stepwise logistic regression analysis confirmed these findings. To establish that this polymorphism is associated with stroke independent of hypertension; we compared stroke patients without hypertension with normotensive controls. Significant difference was observed in genotypic distribution and allelic frequency between the two groups (P = 0.001 and 0.002 respectively). Evaluating the association of this polymorphism with stroke subtypes we found significant associations with cardioembolic stroke (P < 0.001). In conclusion our study suggests that 1347A allele of CYP4F2 gene is an important risk factor for hypertension and ischemic stroke.     

Context-dependent conservation of DNA methyltransferases in bacteria

DNA methytransferases (MTs) in bacteria are best understood in the context of restriction-modification (R-M) systems, which act as bacterial immune systems against incoming DNA including phages, but have also been described as selfish elements. But several orphan MTs, which are not associated with any restriction enzyme, have also been characterized and may protect against parasitism by R-M systems. The occurrence of MTs in these two contexts, namely as part of R-M systems or as orphans, is poorly understood. Here we report the results of a comparative genomic survey of DNA MTs across ～1000 bacterial genomes. We show that orphan MTs overwhelm R-M systems in their occurrence. In general, R-M MTs are poorly conserved, whereas orphans are nearly as conserved within a genus as any average gene. However, oligonucleotide usage and conservation patterns across genera suggest that both forms of MTs might have been horizontally acquired. We suggest that many orphan MTs might be 'degradation' products of R-M systems, based on the properties of orphan MTs encoded adjacent to highly diverged REs. In addition, several fully degraded R-M systems exist in which both the MT and the RE are highly divergent from their corresponding reference R-M pair. Despite their sporadic occurrence, conserved R-M systems are present in strength in two highly transformable genera, in which they may contribute to selection against integration of foreign DNA.