Helical conformations of three crystalline pentapeptide fragments of suzukacillin,a membrane channel forming polypeptide

The crystal structures of three pentapeptide fragments of suzukacillin-A have been determined. Boc-Aib-Pro-Val-Aib-Val-OMe (peptide 1–5) adopts a distorted helical conformation, stabilized by three intramolecular hydrogen bonds (two 5→1, one 4→1). Boc-Ala-Aib-Ala-Aib-Aib-OMe (peptide 6–10) and Boc-Leu-Aib-Pro-Val-Aib-OMe (peptide 16–20) adopt 3₁₀ helical structures stabilized by three and two 4→1 intramolecular hydrogen bonds, respectively. These structures provide substantial support for a largely helical conformation for the suzukacillin membrane channel.     

Temporal variability of forest communities: empirical estimates of population change in 4000 tree species

Long‐term surveys of entire communities of species are needed to measure fluctuations in natural populations and elucidate the mechanisms driving population dynamics and community assembly. We analysed changes in abundance of over 4000 tree species in 12 forests across the world over periods of 6–28 years. Abundance fluctuations in all forests are large and consistent with population dynamics models in which temporal environmental variance plays a central role. At some sites we identify clear environmental drivers, such as fire and drought, that could underlie these patterns, but at other sites there is a need for further research to identify drivers. In addition, cross‐site comparisons showed that abundance fluctuations were smaller at species‐rich sites, consistent with the idea that stable environmental conditions promote higher diversity. Much community ecology theory emphasises demographic variance and niche stabilisation; we encourage the development of theory in which temporal environmental variance plays a central role.     

Interaction of aromatic imino glycoconjugates with jacalin: experimental and computational docking studies

Altering the lectin properties by chemically modified glycoconjugates can have profound effect on their biological applications. In the present case, jacalin has been chosen to study the binding aspects toward glycoconjugates modified by connecting aromatic moieties through imine conjugation at their C-1- or C-2-positions. Out of 10 glycoconjugates, the galactosyl-naphthyl imine (1c) was found to be most effective toward agglutination inhibition (260 times better than galactose), quenching fluorescence intensity, and exhibiting greater binding (K_a, 1.3 × 10⁴ M⁻¹) with jacalin. The specific binding of galactose conjugates and the nonspecific binding of other conjugates have been demonstrated based on ITC. Changes in the secondary structures have been addressed by far- and near-UV CD spectroscopy. The present studies demonstrated that galactose-based conjugates bind at carbohydrate recognition domain (CRD) mainly through polar interactions in addition to exhibiting some nonpolar/hydrophobic interactions, whereas the conjugates other than galactose primarily interact through hydrophobic interactions. Binding of galactosyl conjugates at CRD has been further demonstrated by rigid docking.     

Structural Integrity of the Greek Key Motif in βγ-Crystallins Is Vital for Central Eye Lens Transparency

Background

We highlight an unrecognized physiological role for the Greek key motif, an evolutionarily conserved super-secondary structural topology of the βγ-crystallins. These proteins constitute the bulk of the human eye lens, packed at very high concentrations in a compact, globular, short-range order, generating transparency. Congenital cataract (affecting 400,000 newborns yearly worldwide), associated with 54 mutations in βγ-crystallins, occurs in two major phenotypes nuclear cataract, which blocks the central visual axis, hampering the development of the growing eye and demanding earliest intervention, and the milder peripheral progressive cataract where surgery can wait. In order to understand this phenotypic dichotomy at the molecular level, we have studied the structural and aggregation features of representative mutations.

Methods

Wild type and several representative mutant proteins were cloned, expressed and purified and their secondary and tertiary structural details, as well as structural stability, were compared in solution, using spectroscopy. Their tendencies to aggregate in vitro and in cellulo were also compared. In addition, we analyzed their structural differences by molecular modeling in silico.

Results

Based on their properties, mutants are seen to fall into two classes. Mutants A36P, L45PL54P, R140X, and G165fs display lowered solubility and structural stability, expose several buried residues to the surface, aggregate in vitro and in cellulo, and disturb/distort the Greek key motif. And they are associated with nuclear cataract. In contrast, mutants P24T and R77S, associated with peripheral cataract, behave quite similar to the wild type molecule, and do not affect the Greek key topology.

Conclusion

When a mutation distorts even one of the four Greek key motifs, the protein readily self-aggregates and precipitates, consistent with the phenotype of nuclear cataract, while mutations not affecting the motif display ‘native state aggregation’, leading to peripheral cataract, thus offering a protein structural rationale for the cataract phenotypic dichotomy “distort motif, lose central vision”.     

Development of 5-nitrothiazole derivatives: Identification of leads against both replicative and latent Mycobacterium tuberculosis

Twenty eight 5-nitrothiazole derivatives were synthesized and evaluated for in vitro activities against Mycobacterium tuberculosis (MTB), cytotoxicity against HEK 293T. Among the compounds, 5-nitro-N-(5-nitrothiazol-2-yl)furan-2-carboxamide (20) was found to be the most active compound in vitro with MICs of 5.48 μM against log-phase culture of MTB and also non-toxic up to 100 μM.     

The Mutation V42M Distorts the Compact Packing of the Human Gamma-S-Crystallin Molecule,Resulting in Congenital Cataract

Background

Human γS-crystallin is an important component of the human eye lens nucleus and cortex. The mutation V42M in the molecule causes severe congenital cataract in children. We compare the structure of the mutant protein with that of the wild type in order to understand how structural changes in the mutant relate to the mechanism of opacification.

Methods

Both proteins were made using conventional cloning and expression procedures. Secondary and tertiary structural features of the proteins were analyzed using spectral methods. Structural stabilities of the proteins were analyzed using chemical and thermal denaturation methods. Self-aggregation was monitored using extrinsic spectral probes. Molecular modeling was used to compare the structural features of the two proteins.

Results

While the wild type and mutant have the same secondary structure, molecular modeling and fluorescence analysis suggest the mutant to have a more open tertiary structure, with a larger hydrophobic surface. Experiments using extrinsic probes reveal that the mutant readily self-aggregates, with the suggestion that the aggregates might be similar to amyloidogenic fibrils. Chemical denaturation indicates that while the wild type exhibits the classic two-state transition, V42M goes through an intermediate state, and has a distinctly lower stability than the wild type. The temperature of thermal unfolding of the mutant is also distinctly lower. Further, the mutant readily precipitates and scatters light more easily than the wild type.

Conclusion

The replacement of valine in position 42 by the longer and bulkier methionine in human γS-crystallin perturbs the compact β-sheet core packing topology in the N-terminal domain of the molecule, exposes nonpolar residues thereby increasing the surface hydrophobicity and weakens the stability of the protein, thus promoting self-aggregation leading to light scattering particles. This set of changes in the properties of the mutant offers a molecular insight into the mechanism of opacification.     

Association of tumor necrosis factor alpha, interferon gamma and interleukin 10 gene polymorphisms with peripheral neuropathy in South Indian patients with type 2 diabetes

Diabetic peripheral neuropathy (DPN) is a major global health threat and a common complication of diabetes. Peripheral nerve complications due to irregular cytokine production are eminent factors in many inflammatory diseases. The present study focused on gene polymorphisms of pro and anti-inflammatory cytokines that may be responsible for nerve damage in diabetic neuropathy. We examined three common functional SNPs primarily at the positions on genes of tumor necrosis alpha (TNFα) −308G/A, interferon gamma (IFNγ) +874A/T and interleukin (IL) 10 −1082G/A in order to establish their association with peripheral neuropathy in type 2 diabetes. Results: Genotypic frequencies obtained from TNFα −308G/A gene analysis in DPN group comprised 86.4% of G/A, 10.6% of G/G and 3% of A/A genotype, where as the control group had 94% of G/A, 2% of G/G and 4% of A/A which could not reach the statistical significance with the disease after Bonferroni correction. The IFNγ +874 A/T polymorphism in patient group revealed 33.3% of A/A, 47.5% of A/T and 19.2% of T/T genotype. The A/A genotype had attained statistical significance of P = 0.04 (P corrected); OR 2; 95% CI 1.14–3.64 when compared to controls. The IL10 −1082 G/A polymorphism in the patient group has showed 62.6% of A/A, 21.2% of G/A, 16.2% of G/G genotype, revealing significant association with G/G genotype (P < 0.01, OR 2.9; 95% CI 1.47–5.84) when compared to controls. Conclusion: Our findings indicate that the tested markers within the IFNγ and IL-10 genes, but not the TNFα gene, are significantly associated with peripheral neuropathy in South Indian type 2 diabetic patients. The study shows that the ‘high-producer’ IL-10 −1082 G/G genotype and the ‘low-producer’ IFNγ +874 A/A genotype may be responsible for the down regulation of immune response leading to inflammation in this setting.     

Ntlim1, a PAL-box binding factor,controls promoter activity of the horseradish wound-inducible peroxidase gene

To understand molecular mechanisms underlying wound-induced expression of plant peroxidase genes, the promoter of a horseradish C2 peroxidase (prxC2) gene was analyzed. We had previously isolated a tobacco nuclear protein, Ntlim1, as a trans factor binding to a PAL-box motif of the prxC2 promoter; however, the function of the Ntlim1 trans factor and the PAL-box motif in wound-responsive expression of the prxC2 gene remains unclear. Here, we found that the prxC2 promoter without the intact PAL-box motif failed to direct a normal level of both the basal and the wound-induced expression of -glucuronidase (GUS) reporter gene in transgenic tobacco plants, indicating that the PAL-box motif functions as an essential cis element of the prxC2 promoter. We also found that antisense expression of Ntlim1 in transgenic plants carrying the prxC2 promoter::GUS chimeric construct decreased not only the level of the basal and the wound-induced expression of the GUSreporter gene but also the extent of wound inducibility of the prxC2 promoter itself. This result indicates that Ntlim1 is required for the basal level of prxC2 promoter activity as well as its up-regulation under wound stress. Moreover, consistent with the results obtained in planta, result from super-shift assay indicates that the Ntlim1 binds to the PAL-box motif independently of wound stress.