Abstract:
The H1-NMR spectroscopy was a suitable method for the determination of glycine betaine in a halotolerant cyanobacterium, A.halophytica. The amount of glycine betaine from A.halophytica was 9.7 nmol/10 6 cells when the cells were grown in non-salt stressed condition (0.5M NaCl). Glycine betaine accumulation increased up to 8 folds when A.halophytica cells were cultivated in salt stressed condition (2M NaCl). Betaine aldehyde dehydrogenase was purified about 18-fold from A.halophytica with a specific activity of 290.8 micro mol min -1mg-1. The procedure included fractional precipitation with 35-70% ammonium sulfate and DEAE-cellulose column chromatography. The optimum condition for betaine aldehyde dehydrogenase activity was pH 7.5 and 25 ํC. The A.halophytica betaine aldehyde dehydrogenase was specific for betaine aldehyde and NAD+ with a Km of 91 and 71.4 micro M respectively. The Vmax of betaine aldehyde dehydrogenase was 175.4 micro mol min -1mg-1. The acetaldehyde and ethanolamine as substrate analogs showed strong inhibition towards betaine aldehyde dehydrogenase activity. NaCl and KCl at or below 0.1 M stimulated BADH activity. At higher than 0.1 M KCl the enzyme activity declined and returned to the control level whereas for higher than 0.1 M NaCl the enzyme activity was inhibited. CaCl2 and MgCl2 at all concentrations tested up to 0.5 M inhibited betaine aldehyde dehydrogenase activity. The p-chloromercuriphenyl sulfonic acid was a potent inhibitor of the enzyme. Preincubation of the enzyme with DTT could protect the enzyme activity against the inhibition by p-chloromercuriphenyl sulfonic acid. Gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis suggested that the molecular weight of the enzyme was 120,000 dalton and it was likely that the enzyme was a tetramer of 30,000 dalton subunits. The relationship between salt stress and betaine aldehyde dehydrogenase activity revealed that the high external salinity increased specific activity of betaine aldehyde dehydrogenase. The data supported the idea that the synthesis of betaine aldehyde dehydrogenase could be induced by external salinization.
