Ganesan Saibaba

Optimization of polyhydroxybutyrate production by marine Bacillus megaterium MSBN04 under solid state culture

A marine sponge-associated bacterium Bacillus megaterium MSBN04 was used for the production of polyhydroxybutyrate (PHB) under solid state culture (SSC). A central composite design (CCD) was employed to optimize the production medium and to find out the interactive effects of four independent variables, viz. tapioca industry waste, palm jaggery, horse gram flour and trace element solution on PHB production. The maximum yield of PHB 8.637 mg g(-1) of substrate (tapioca industry waste) was achieved from biomass 15.203 mg g(-1) of substrate, using statistically optimized medium. The horse gram flour (nitrogen source) and trace element solution were found to be critical control factors for PHB synthesis. The (1)H NMR analysis revealed that the polymer was a PHB monomer. PHB obtained from this study having high molecular weight (6.7×10(5) Da) with low polydispersity index (PDI) value (1.71) and produced PHB was used to synthesize PHB polymeric nanoparticles using solvent displacement approach. Therefore, B. megaterium MSBN04 is an ideal candidate that can be exploited biotechnologically for the commercial production of PHB under solid state culture.

A statistical approach for optimization of polyhydroxybutyrate production by marine Bacillus subtilis MSBN17

The important biological macromolecule polyhydroxybutyrate (PHB) producing Bacillus subtilis was isolated from the marine sponge Callyspongia diffusa and identified by means of 16S rRNA analysis. The central composite design (CCD) was used to optimize the PHB production using cheap raw materials such as pulp industry waste (PIW), tamarind kernel powder (TKP), palm jaggery (PJ) and green gram flour (GGF). The extracted polymer was characterized by (1)H NMR analysis. The PIW was fed at three different intervals and the maximum production of PHB (19.08g/L) was attained after a period of 40h of incubation of B. subtilis. Dissolved oxygen, sodium chloride and nitrogen source were found to be the critical control factors that affected the PHB polymer production. The present investigation demonstrates an inexpensive model of producing PHB green thermoplastics in vitro for biomedical applications.

Synthesis of carbohydrate polymer encrusted gold nanoparticles using bacterial exopolysaccharide: a novel and greener approach

In the present study, a marine sponge-associated endosymbiotic bacterium Bacillus megaterium MSBN04 was evaluated for exopolysaccharide (EPS) production. The production process was optimized by central composite design (CCD). The productivity was increased up to 5.62 g L−1 with sucrose as sole carbon source. The secreted EPS was characterized by NMR analysis, confirming the presence of monosaccharide units such as α-D-glucose and α-D-galactose, which further confirms that the secreted EPS is a heteropolysaccharide. The purified EPS showed considerable flocculating activity (45.41%) with 4 mg L−1 of EPS. Using EPS as reducing and stabilizing agent, gold nanoparticles (AuNPs) were synthesized. The synthesized AuNPs (5–20 nm) were of spherical crystalline nature and capped with an EPS layer and were characterized by transmission electron microscopy (TEM) and X-ray diffraction (XRD) analysis. The synthesis of AuNPs was dependent on the concentration of EPS. The synthesized AuNPs showed significant antibacterial activity against clinical pathogenic bacteria. Hence, EPS-mediated synthesis of AuNPs is an alternative approach to chemical synthesis and thus it is an environmentally benign, greener and economical approach.

Exploration of salivary proteins in buffalo: an approach to find marker proteins for estrus

Saliva is considered as the best source of biological material for biomarker discovery studies since it is noninvasive in comparison to other body sources. Usually buffalo cannot precisely express estrus signals. Hence, there is a need for concise methods to detect the time of estrus to ensure the success of artificial insemination. Therefore, we have established a reference proteome map on the whole saliva of buffalo during their estrous cycle with special reference to estrus. Nearly 12 bands have been observed using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) of whole saliva. Collectively, 179 proteins are identified with respect to different phases of the estrous cycle using mass spectrometry. On the whole, 37 proteins are exclusively expressed in the estrus phase, which include β‐enolase, Toll‐like receptor (TLR) 4, clusterin, lactoperoxidase, serotransferrin, TGM3, UBA6, and transducin. Among the proteins, β‐enolase and TLR 4 were validated, and their specific expression was found during estrus as compared to other phases using immunoblot. The functional annotation reveals many as binding proteins in the estrus saliva when compared to the other phases. The present findings conclude that the proteomic approach adopted to identify the proteins from buffalo saliva around the estrous cycle may provide a new tool for screening the estrus phase. The results further conclude that the specific expression of β‐enolase and TLR 4 can be taken as the indicator of estrus in buffalo.—Muthukumar, S., Rajkumar, R., Rajesh, D., Saibaba, G., Liao, C.‐C., Archunan, G., Padmanabhan, P., Gulyas, B., Exploration of salivary proteins in buffalo: an approach to find marker proteins for estrus. FASEB J. 28, 4700–4709 (2014). www.fasebj.org

Urinary Lipocalin Protein in a Female Rodent with Correlation to Phases in the Estrous Cycle: An Experimental Study Accompanied by In Silico Analysis

Male urinary lipocalin family proteins, practically odorant-binding proteins but also could be pheromones by themselves, in rodents act as a shuttle for chemosignal communication and facilitate delivery of the signals for access to congeners. However, presence of this protein in urine of female rodents has not yet been reported. Therefore, the present investigation was carried out to find if lipocalin family protein is present in the urine of female house rat and, if so, to find whether its expression differs between the phases in the estrous cycle. The rat urinary protein was separated in single dimensional gel electrophoresis. A 14.5 kDa lipocalin protein appeared in the urine prominently during the estrus and metestrus phases compared to proestrus and diestrus phases. The expression of this protein in the urine was very low in ovariectomized rats. MALDI-TOF/MS analysis affirmed the 14.5 kDa protein as a lipocalin family protein. Analysis adopting bio-informatics tools further proved the protein as a lipocalin family member. Thus, this study for the first time demonstrated the presence of a lipocalin family protein in the urine of a female rodent and it was highly expressed during estrus phase. This lipocalin protein in female rat urine may facilitate a chemosignal function independently of a pheromone or in association with a specific pheromone.

Characterization of salivary protein during ovulatory phase of menstrual cycle through MALDI-TOF/MS

CONTEXT Predicting ovulation is the basis on which the fertile period is determined. Nowadays there are many methods available to detect the ovulatory period. Unfortunately, these methods are not always effective for accurate detection of ovulation. Hence, an attempt was made to detect ovulation through single dimension sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of protein with the help of saliva ferning. AIMS The aim of this study was to determine the association of protein level with endogenous reproductive hormone level across the menstrual cycle. SETTINGS AND DESIGN Salivary protein and its confirmation were evaluated during menstrual cycle followed by SDS-PAGE and Mass spectrometry. STATISTICAL METHOD USED: The protein content present in saliva throughout menstrual cycle is trail by SPSS statistical software version. MATERIALS AND METHODS Salivary proteins were investigated serially during pre-ovulatory, ovulatory and post-ovulatory periods of normal menstrual cycle in eighteen healthy volunteers. The samples were collected in three consecutive menstrual cycles. Salivary protein was estimated and analyzed by single dimension SDS-PAGE. RESULTS The results revealed significant variations in protein concentrations during the menstrual cycle. Protein levels were maximum during ovulation and minimum during postovulatory phase. Further, single dimension SDS-PAGE analysis showed seven different fractions of proteins is from 14-90 kilo Dalton (kDa) in the three phases of the menstrual cycle. CONCLUSIONS Among the proteins, 48 kDa protein was more predominantly exhibited during ovulatory phase than pre and post-ovulatory phase. The present study indicates that the protein level and the specific protein band (48 kDa) through MALDI-TOF MS analysis might serve as an indicator for ovulation.

Marine sponge-associated bacteria as a potential source for polyhydroxyalkanoates

Abstract Marine sponges are filter feeding porous animals and usually harbor a remarkable array of microorganisms in their mesohyl tissues as transient and resident endosymbionts. The marine sponge-microbial interactions are highly complex and, in some cases, the relationships are thought to be truly symbiotic or mutualistic rather than temporary associations resulting from sponge filter-feeding activity. The marine sponge-associated bacteria are fascinating source for various biomolecules that are of potential interest to several biotechnological industries. In recent times, a particular attention has been devoted to bacterial biopolymer (polyesters) such as intracellular polyhydroxyalkanoates (PHAs) produced by sponge-associated bacteria. Bacterial PHAs act as an internal reserve for carbon and energy and also are a tremendous alternative for fossil fuel-based polymers mainly due to their eco-friendliness. In addition, PHAs are produced when the microorganisms are under stressful conditions and this biopolymer synthesis might be exhibited as one of the survival mechanisms of sponge-associated or endosymbiotic bacteria which exist in a highly competitive and stressful sponge-mesohyl microenvironment. In this review, we have emphasized the industrial prospects of marine bacteria for the commercial production of PHAs and special importance has been given to marine sponge-associated bacteria as a potential resource for PHAs.

Structural elucidation of estrus urinary lipocalin protein (EULP) and evaluating binding affinity with pheromones using molecular docking and fluorescence study

Transportation of pheromones bound with carrier proteins belonging to lipocalin superfamily is known to prolong chemo-signal communication between individuals belonging to the same species. Members of lipocalin family (MLF) proteins have three structurally conserved motifs for delivery of hydrophobic molecules to the specific recognizer. However, computational analyses are critically required to validate and emphasize the sequence and structural annotation of MLF. This study focused to elucidate the evolution, structural documentation, stability and binding efficiency of estrus urinary lipocalin protein (EULP) with endogenous pheromones adopting in-silico and fluorescence study. The results revealed that: (i) EULP perhaps originated from fatty acid binding protein (FABP) revealed in evolutionary analysis; (ii) Dynamic simulation study shows that EULP is highly stable at below 0.45 Å of root mean square deviation (RMSD); (iii) Docking evaluation shows that EULP has higher binding energy with farnesol and 2-iso-butyl-3-methoxypyrazine (IBMP) than 2-naphthol; and (iv) Competitive binding and quenching assay revealed that purified EULP has good binding interaction with farnesol. Both, In-silico and experimental studies showed that EULP is an efficient binding partner to pheromones. The present study provides impetus to create a point mutation for increasing longevity of EULP to develop pheromone trap for rodent pest management.

Proteomic analysis of human saliva: An approach to find the marker protein for ovulation

Human saliva contains numerous molecules that play a variety of roles. Among them there are proteins which serve as biomarkers of various physiological and/or pathological conditions. Compared to other body fluids, saliva is the most convenient material for investigations, and especially for monitoring the disease conditions. Presently, there is an increasing need to develop a noninvasive method to identify the time of ovulation in humans to ensure successful fertilization, and for evolving strategies for family planning. The present investigation has been an attempt to identify one or more proteins in the human saliva that would be an indicator(s) of ovulation. SDS-PAGE of salivary proteins showed seven prominent bands during the different phases of the menstrual cycle. Particularly, the 14.5kDa band was highly expressed during the ovulatory phase. Eleven proteins were identified in this band of which ten were highly specific to the ovulatory phase. Among those proteins the intense expression of Cystatin-S was validated using immunoblot analysis (p<0.05). The functional annotation of salivary proteins revealed a high percentage of proteins that engage in binding and regulatory activities. The present results indicate that salivary proteins, particularly those present during the ovulatory phase, might be used as biomarkers for impending ovulation.

Does salivary protein(s) act an ovulation indicator for women?: A hypothesis

Ovulation is an important physiological process in human, and its effect may reflect in body fluids via secretion of biomolecules such as proteins, amino acids, antioxidants, antimicrobial peptides and so on. Recently, the non-invasive sampling approaches are used to diagnose disease status and access health condition of human. Saliva comprises various proteins which are secreted through salivary glands. The proteins present in the saliva may vary in their expression according to the hormonal level and physiological nature of the body which are said to be hormone receptors, stress proteins and antimicrobial peptides. Therefore, it is postulated that saliva can be used in the detection of ovulation time in human using specific protein(s) expression and which can be considered as a best non-invasive method. The identification of these proteins by adopting LC-MS/MS followed by Western blot analysis are possible to identify a promising biomarker for ovulation detection in human.