Showing papers by "Sathyanarayana N. Gummadi published in 2011"

Metabolism of glucose and xylose as single and mixed feed in Debaryomyces nepalensis NCYC 3413: production of industrially important metabolites

Abstract: Efficient conversion of hexose and pentose (glucose and xylose) by a single strain is a very important factor for the production of industrially important metabolites using lignocellulose as the substrate. The kinetics of growth and polyol production by Debaryomyces nepalensis NCYC 3413 was studied under single and mixed substrate conditions. In the presence of glucose, the strain produced ethanol (35.8 ± 2.3 g/l), glycerol (9.0 ± 0.2 g/l), and arabitol (6.3 ± 0.2 g/l). In the presence of xylose, the strain produced xylitol (38 ± 1.8 g/l) and glycerol (18 ± 1.0 g/l) as major metabolites. Diauxic growth was observed when the strain was grown with different combinations of glucose/xylose, and glucose was the preferred substrate. The presence of glucose enhanced the conversion of xylose to xylitol. By feeding a mixture of glucose at 100 g/l and xylose at 100 g/l, it was found that the strain produced a maximum of 72 ± 3 g/l of xylitol. A study of important enzymes involved in the synthesis of xylitol (xylose reductase (XR) and xylitol dehydrogenase (XDH)), glycerol (glycerol-3-phosphate dehydrogenase (G3PDH)) and ethanol (alcohol dehydrogenase (ADH)) in cells grown in the presence of glucose and xylose revealed high specific activity of G3PDH and ADH in cells grown in the presence of glucose, whereas high specific activity of XR, XDH, and G3PDH was observed in cells grown in the presence of xylose. To our knowledge, this is the first study to elaborate the glucose and xylose metabolic pathway in this yeast strain.

Synthesis of both enantiomers of ethyl-4-chloro-3-hydroxbutanoate from a prochiral ketone using Candida parapsilosis ATCC 7330

Abstract: Candida parapsilosis ATCC 7330 when grown in a medium containing glycerol reduced ethyl-4-chloro-3-oxobutanoate to ( R )-ethyl-4-chloro-3-hydroxybutanote (ee >99%, yield: 94%) while glucose and sucrose grown cells yielded ( S )-ethyl-4-chloro-3-hydroxybutanote (ee >99%, yield: 96%). The activity of ethyl-4-chloro-3-oxobutanoate reductase was higher in glucose-grown cells (160 U/g protein) when compared to sucrose (158 U/g protein) and glycerol (22 U/g protein). Both the enantiomers of ethyl-4-chloro-3-hydroxybutanoate (ee >99%) can thus be obtained using Candida parapsilosis ATCC 7330 by altering the carbon source in the growth medium.

Candida parapsilosis ATCC 7330 can also deracemise 1-arylethanols

Abstract: Candida parapsilosis ATCC 7330 grown using different culture conditions (inoculum size 4% (v/v), inoculum age 12 h, and harvest time 14 h) from those previously reported (inoculum size 2% (v/v), inoculum age 24 h, and harvest time 44 h) successfully deracemised racemic 1-arylethanols and 4-phenyl-2-butanol to the (R)-enantiomer (ee up to >99%). The deracemisation of racemic 1-aryl ethanol proceeds via (i) enantioselective oxidation of (S)-enantiomer followed by (ii) reduction of the ketone formed to give the racemic alcohol which gets kinetically resolved thus enriching for the (R)-enantiomer from the racemate. This is the first report on the deracemisation of 1-arylethanols using Candida parapsilosis ATTC 7330 via dynamic kinetic resolution.

Differential response of the catalase, superoxide dismutase and glycerol-3-phosphate dehydrogenase to different environmental stresses in Debaryomyces nepalensis NCYC 3413

Abstract: The effect of salt, pH, and temperature stress on the cellular level of antioxidant enzymes, catalase and superoxide dismutase (SOD) and glycerol-3-phosphate dehydrogenase (G3PDH) was studied in Debaryomyces nepalensis NCYC 3413, a halotolerant yeast. The catalase activity increased in different phases, while SOD and G3PDH activities declined in late stationary phase. A significant increase in SOD activity was observed under different stress as compared to control. Salt and temperature stress enhanced the catalase activity where as it was suppressed by pH stress. G3PDH level increased with salt stress, however, no significant change was observed under pH and temperature stress. The observations recorded in this investigation suggested that D. nepalensis has an efficient protective mechanism of antioxidant enzymes and G3PDH against salt, pH, and temperature stresses.

Enhanced degradation of caffeine and caffeine demethylase production by Pseudomonas sp. in bioreactors under fed-batch mode.

Abstract: The growth of Pseudomonas sp was studied in fed-batch process with an aim to improve the caffeine degradation rate and caffeine demethylase activity The effects of varying initial caffeine concentrations in the batch mode, increase in the number of feeds, varying feed flow rates, and added nutrients to the feed on the fed-batch process were investigated A maximum caffeine degradation rate of 082 g/L h and maximum caffeine demethylase activity of 26 U/mg were achieved using manual intermittent pulse feeds of caffeine with substrate concentration as feedback parameter for the fed batch started with an initial caffeine concentration of 3 g/L A slight increase in the caffeine degradation rate (085 g/L h) and caffeine demethylase activity (34 U/mg) was observed when the additional nutrients were added along with caffeine in the feed This is the first report showing complete degradation of large magnitudes of caffeine amounting to 237 g in 75 h These results show that the fed-batch conditions achieved in this study using Pseudomonas sp facilitate the development of a sustainable bioprocess to degrade the high concentrations of caffeine in industrial effluents

Flip-flop of phospholipids in proteoliposomes reconstituted from detergent extract of chloroplast membranes: kinetics and phospholipid specificity.

Abstract: Eukaryotic cells are compartmentalized into distinct sub-cellular organelles by lipid bilayers, which are known to be involved in numerous cellular processes. The wide repertoire of lipids, synthesized in the biogenic membranes like the endoplasmic reticulum and bacterial cytoplasmic membranes are initially localized in the cytosolic leaflet and some of these lipids have to be translocated to the exoplasmic leaflet for membrane biogenesis and uniform growth. It is known that phospholipid (PL) translocation in biogenic membranes is mediated by specific membrane proteins which occur in a rapid, bi-directional fashion without metabolic energy requirement and with no specificity to PL head group. A recent study reported the existence of biogenic membrane flippases in plants and that the mechanism of plant membrane biogenesis was similar to that found in animals. In this study, we demonstrate for the first time ATP independent and ATP dependent flippase activity in chloroplast membranes of plants. For this, we generated proteoliposomes from Triton X-100 extract of intact chloroplast, envelope membrane and thylakoid isolated from spinach leaves and assayed for flippase activity using fluorescent labeled phospholipids. Half-life time of flipping was found to be 6±1 min. We also show that: (a) intact chloroplast and envelope membrane reconstituted proteoliposomes can flip fluorescent labeled analogs of phosphatidylcholine in ATP independent manner, (b) envelope membrane and thylakoid reconstituted proteoliposomes can flip phosphatidylglycerol in ATP dependent manner, (c) Biogenic membrane ATP independent PC flipping activity is protein mediated and (d) the kinetics of PC translocation gets affected differently upon treatment with protease and protein modifying reagents.

Use of Response Surface Methodology to Study the Combined Effect of Salt, pH, and Temperature on the Growth of Food-Spoiling Halotolerant Yeast, Debaryomyces nepalensis NCYC 3413

Abstract: This paper reports the interaction of salt (NaCl and KCl), initial pH, and temperature and their effects on the specific growth rate and lag phase of food spoiling halotolerant yeast, Debaryomyces nepalensis. The optimization of salt, initial pH, and temperature was carried out using response surface methodology based on central composite design. The mathematical model showed that salt has a significant effect on specific growth rate and lag phase of D. nepalensis. The optimal conditions of salt concentration, pH, and temperature of growth were found to be 0.3 M NaCl, 7.1, 26 °C and 0.6 M KCl, 5.6, 25°C, respectively. Under these conditions, a maximum specific growth rate of 0.41 and 0.5 h−1 was observed in medium containing NaCl and KCl, respectively. Lag phase can be increased most effectively either by increasing salt concentrations or both by decreasing (≤20°C) or increasing the temperature (≥40°C) with moderate (1.5 M) or low salt concentration (0.5 M), respectively. Results show that D. nepalensis need to generate weak acids to maintain the intracellular pH under pH and saline stress conditions. The results obtained in this study will be helpful in using optimal conditions for the maximum growth of the strain for the production of certain metabolites like organic acids and glycerol and designing food preservation procedures.