ABO blood groups and fertility—with special reference to intrauterine selection due to materno-fetal incompatibility

The purpose of the present investigation was to study whether there is differential fertility between different mating types of ABO blood group system. Selective force which is operating through maternal-fetal incompatibility has been observed in the differential fertility between compatible and incompatible mating groups in the present sample of 183 families of Visakhapatnam town of Andhra Pradesh, India. The differences in the mean numbers of pregnancies as well as living children between the two major mating groups, compatible and incompatible are significant. The fertility rates of O fathers and O mothers were significantly higher than those in matings in which neither parents belongs to O. The selection is operating to reduce the gene ratio of A and to increase the gene ratios of O and B in this sample.     

Identification of amino acid residues at the active site of human liver serine hydroxymethyltransferase

Chemical modification of amino acid residues with phenylglyoxal, diethylpyrocarbonate, and N-bromosuccinimide indicated that at least one residue each of arginine, histidine, and tryptophan were necessary for the activity of human liver serine hydroxymethyltransferase. Protection by substrates suggested that these residues might occur at the active site of the enzyme.     

Changes in leucyl-tRNAs and aminoacyl tRNA synthetases in developing and aging soybean cotyledons

Leucine specific tRNA of soybean cotyledons was frationated into six peaks (1–6). The relative amounts of Leu-tRNA 5 and 6 are lower in developing cotyledons than in germinating cotyledons. Leu-tRNA synthetase from developing cotyledons is less active in aminoacylating Leu-tRNA 5 and 6 compared to enzyme from 5-day-old germinating cotyledons. Leu-tRNA synthetase from cotyledons of germinating seedlings and developing cotyledons can be fractionated into three peaks (1–3). Peak 1 in the developing cotyledon is about 36% less than peak 1 from 5-day-old germinating cotyledons. Peaks 2 and 3 from developing cotyledons are about 10 and 18% higher than from germinating cotyledons, respectively. Peak 1 from developing cotyledons acylates all six species of Leu-tRNA in contrast with peak 1 from germinating cotyledons, which essentially acylates only Leu-tRNA 5 and 6. The specificity of peaks 2 and 3 towards Leu-tRNA 1–4 is identical in both the organs.     

Plants fabricate Fe-nanocomplexes at root surface to counter and phytostabilize excess ionic Fe

While evaluating the impact of iron nanoparticles (NPs) on terrestrial plants we realized potential of root system of intact plants to form orange–brown complexes constituted of NPs around their roots and at bottom/side of tubes when exposed to FeCl₃. These orange–brown complexes/plaques seen around roots were similar to that reported in wetland plants under iron toxicity. Transmission electron microscopy coupled with energy dispersive X-ray analysis revealed that orange–brown complexes/plaques, formed by root system of all 16 plant species from 11 distinct families tested, were constituted of NPs containing Fe. Selected area electron diffraction and powder X-ray diffraction spectra showed their amorphous nature. Thermogravimetric and fourier transform infra-red analysis showed that these Fe-NPs/nanocomplexes were composed of iron-oxyhydroxide. These plant species generated orange–brown Fe-NPs/nanocomplexes even under strict sterile conditions establishing inbuilt and independent potential of their root system to generate Fe-NPs. Root system of intact plants showed ferric chelate reductase activity responsible for reduction of Fe³⁺ to Fe²⁺. Reduction of potassium ferricyanide by root system of intact plants confirmed that root surface possess strong reducing strength, which could have played critical role in reduction of Fe³⁺ and formation of Fe-NPs/nanocomplexes. Atomic absorption spectrophotometric analysis revealed that majority of iron was retained in Fe-nanocomplexes/plaques, while only 2–3 % was transferred to shoots, indicating formation of nanocomplexes is a phytostabilization mechanism evolved by plants to restrict uptake of iron above threshold levels. We believe that formation of Fe-NPs/nanocomplexes is an ideal homeostasis mechanism evolved by plants to modulate uptake of desired levels of ionic Fe.     

In vitro removal of human IgG autoantibodies by affinity filtration using immobilized L-histidine onto PEVA hollow fiber membranes

Histidine was immobilized onto PEVA membrane to obtain an affinity support for human IgG removal from serum with a view to clinical apheresis for the treatment of autoimmune diseases. These membranes were able to remove in vitro several autoantibodies from the serum of SLE patients.     

Mutagenesis of Varicella-Zoster Virus Glycoprotein B: Putative Fusion Loop Residues Are Essential for Viral Replication,and the Furin Cleavage Motif Contributes to Pathogenesis in Skin Tissue In Vivo

Glycoprotein B (gB), the most conserved protein in the family Herpesviridae, is essential for the fusion of viral and cellular membranes. Information about varicella-zoster virus (VZV) gB is limited, but homology modeling showed that the structure of VZV gB was similar to that of herpes simplex virus (HSV) gB, including the putative fusion loops. In contrast to HSV gB, VZV gB had a furin recognition motif ([R]-X-[KR]-R-|-X, where | indicates the position at which the polypeptide is cleaved) at residues 491 to 494, thought to be required for gB cleavage into two polypeptides. To investigate their contribution, the putative primary fusion loop or the furin recognition motif was mutated in expression constructs and in the context of the VZV genome. Substitutions in the primary loop, W180G and Y185G, plus the deletion mutation Δ⁴⁹¹RSRR⁴⁹⁴ and point mutation ⁴⁹¹GSGG⁴⁹⁴ in the furin recognition motif did not affect gB expression or cellular localization in transfected cells. Infectious VZV was recovered from parental Oka (pOka)-bacterial artificial chromosomes that had either the Δ⁴⁹¹RSRR⁴⁹⁴ or ⁴⁹¹GSGG⁴⁹⁴ mutation but not the point mutations W180G and Y185G, demonstrating that residues in the primary loop of gB were essential but gB cleavage was not required for VZV replication in vitro. Virion morphology, protein localization, plaque size, and replication were unaffected for the pOka-gBΔ⁴⁹¹RSRR⁴⁹⁴ or pOka-gB⁴⁹¹GSGG⁴⁹⁴ virus compared to pOka in vitro. However, deletion of the furin recognition motif caused attenuation of VZV replication in human skin xenografts in vivo. This is the first evidence that cleavage of a herpesvirus fusion protein contributes to viral pathogenesis in vivo, as seen for fusion proteins in other virus families.Varicella-zoster virus (VZV), an alphaherpesvirus, causes chicken pox (varicella) as a primary infection and shingles (zoster) upon reactivation from infected ganglia in humans (reviewed in reference 16). Although not yet investigated in VZV, herpesvirus entry requires fusion of the virus envelope with cell membranes governed by viral glycoprotein B (gB) and gH/gL, which are conserved across the family Herpesviridae (12, 27, 57). gB is the most conserved glycoprotein, with its function as a fusion protein well documented for several of the herpesviruses (10, 19, 38, 48, 51, 52).Open reading frame 31 (ORF31) codes for the 931 amino acids of VZV gB (18, 37). The successive N- and O-linked glycosylation plus the sialation and sulfation of VZV gB yields a mature protein with a molecular mass of approximately 140 kDa (45). Upon maturation, gB is cleaved, presumably by cellular proteases, into two polypeptides of 66 and 68 kDa. Intracellular trafficking of gB was shown to be dependent upon amino acid motifs in the cytoplasmic domain (24-26). In transfection studies, gB was transported to the cellular surface where it was endocytosed and localized to the trans-Golgi, where envelopment of viral particles is thought to occur.The structures of gB in the two human alphaherpesviruses, VZV and herpes simplex virus type 1 (HSV-1), are likely to be very similar as they have 49% amino acid identity (reviewed in reference 16). The ectodomain of HSV-1 gB was shown to form a spike that consisted of trimers with the structural homology to gG of vesicular stomatitis virus (28). Heldwein et al. (28) proposed that HSV-1 gB is a class II fusion protein based on homology to VSV G. The herpesvirus gB monomer was divided into five domains, I to V. Domain I consisted of a continuous amino acid sequence that folded into a pleckstrin homology-like domain, while domain II was comprised of two discontinuous segments, which also had a pleckstrin homology-like domain. A loop region exposed to the exterior of gB connected domain II with domain III. Domain III was comprised of three discontinuous segments and connected to the external loop by a long α helix that ended in a central coiled coil. Domain IV crowned gB and was connected to domain V, which stretched from the top to the bottom of the gB monomer, forming the core of the trimer making contacts with the two other subunits. The structural homology and lack of furin cleavage suggest that the herpesvirus gB and VSV G proteins have undergone convergent evolution.Although not proven experimentally, VZV gB is likely to be cleaved by the subtilisin-like proprotein convertase furin as the glycoprotein has a furin recognition motif [R]-X-[KR]-R-|-X (where | indicates the position at which the polypeptide is cleaved) (29). The [R]-X-[KR]-R-|-X motif is conserved in gBs for all of the herpesvirus families (5, 9, 21, 36, 40, 53, 63, 64). This site has been shown to be dispensable for the replication of human cytomegalovirus (HCMV), bovine herpesvirus type 1 (BHV-1), and pseudorabies virus (PRV) in vitro (32, 49, 58). Furin site mutants for BHV-1 and PRV show an altered phenotype in vitro, but effects were not examined in vivo. HSV-1 gB is not cleaved and lacks the [R]-X-[KR]-R-|-X motif at the canonical site, which is of interest because HSV-1 is genetically the most closely related human herpesvirus to VZV.Domain I of HSV gB showed structural conservation of putative fusion loops similar to those found in domain IV of the VSV G protein (28). Despite the lack of conserved amino acids within these loops, the hydrophobicity of the residues appears to be conserved for the Herpesviridae (4). Substitution of hydrophobic residues in Epstein-Barr virus gB and linker insertion mutagenesis close to the putative fusion loops of HSV-1 gB abrogated fusion based on in vitro transfection studies (4, 22, 34). However, the effect of substitutions in these putative fusion loops on viral replication has not been characterized. Since the development of fusion assays for VZV has proven elusive, the effect of substitutions in the putative fusion loop using viral mutagenesis to make recombinant viruses provides an alternative approach for identifying functional residues in VZV gB.In contrast to HSV-1, VZV is a human-restricted pathogen (reviewed in reference 16). To study the pathogenesis of VZV in vivo, well-established human xenograft models have been developed using SCID mice (6, 7, 13, 14, 41, 44, 54, 65). Lesions formed by VZV in the skin are similar to those seen in human subjects following primary infection (15, 43). The relevance of the model was demonstrated by studies with the varicella vaccine virus (vOka) that exhibited decreased growth in skin xenografts in vivo but does not cause disease in the healthy human host. In contrast, the vaccine virus and its parent strain, parental Oka (pOka), have indistinguishable replication kinetics in vitro (15, 43).The present study was designed to investigate the effects of structure-based targeted mutations in VZV gB on viral replication in cultured cells and in human skin xenografts in the SCIDhu mouse model. This was performed in the context of infectious virus recovered using the self-excisable bacterial artificial chromosome (BAC) containing the genome of a clinical isolate, Oka (62). The roles of the conserved residues W180 (gB-W180G) and Y185 (gB-Y185G) in the putative fusion loop were evaluated using glycine substitution, and the role of the furin recognition motif (⁴⁹¹RSRR⁴⁹⁴) was assessed by a complete deletion of the furin motif (gBΔ⁴⁹¹RSRR⁴⁹⁴) or a substitution of the arginine residues with glycine (gB⁴⁹¹GSGG⁴⁹⁴) to conserve the carbon backbone.     

Bioconversion of isoflavone glycosides to aglycones,mineral bioavailability and vitamin B complex in fermented soymilk by probiotic bacteria and yeast

Aim: To study the role of β‐glucosidase producing probiotic bacteria and yeast in the biotransformation of isoflavone glycosides to aglycones, mineral bioavailability and vitamin B complex in fermented soymilk. Methods and Results: Five isolates of probiotic lactic acid bacteria (LAB), Lactobacillus acidophilus B4496, Lactobacillus bulgaricus CFR2028, Lactobacillus casei B1922, Lactobacillus plantarum B4495 and Lactobacillus fermentum B4655 with yeast Saccharomyces boulardii were used to ferment soymilk to obtain the bioactive isoflavones, genistein and daidzein. High‐performance liquid chromatography was used to analyse the concentration of isoflavones. Bioactive aglycones genistein and daidzein after 24 and 48 h of fermentation ranged from 97·49 to 98·49% and 62·71 to 92·31% respectively with different combinations of LAB with yeast. Increase in bioavailability of minerals and vitamin B complex were also observed in fermented soymilk. Conclusions: LAB in combination with yeast S. boulardii has great potential for the enrichment of bioactive isoflavones, enhancing the viability of LAB strains, decreasing the antinutrient phytic acid and increasing the mineral bioavailability in soymilk fermentation. Significance and Impact of the Study: Fermentation of soymilk with probiotic organisms improves the bioavailability of isoflavones, assists in digestion of protein, provides more soluble calcium, enhances intestinal health and supports immune system. Increased isoflavone aglycone content in fermented soymilk improves the biological functionality of soymilk.     

Natural and chemically induced oligomeric ribonucleases: structural study by immobilized metal ion affinity electrophoresis and their functional relationship

Oligomerization can endow proteins with novel structural and catalytic properties. The native dimer of bovine seminal ribonucleases (BS-RNase) binds, melts and catalyses the hydrolysis of double-stranded ribonucleic acids 30-fold better than its pancreatic homologue, the monomeric RNase A. Chemically induced oligomers of pancreatic RNase A are also found to show an increase in enzyme activity on double-stranded poly(A).poly(U) (Libonati, M. Bertoldi, M. and Sorrentino, S. (1996) Biochem. J. 318, 287-290) and, therefore, can be considered as potential immunosuppressive and cytotoxic agents. We report here a study on the relationship between surface histidine topography in oligomeric forms of these ribonucleases and their catalytic properties. Subtle changes in structure conformation of both BS-RNase and oligomeric RNase A are shown to result in a modification of the affinity of these proteins toward the immobilized transition-metal chelate, IDA-Cu(II). Because, such conformational change has been shown to correlate with an improvement of the newly acquired biological activities upon oligomerization, we can conclude that surface histidines topography constitutes an exquisite probe for the study of protein structure/function relationship.