Purification and properties of sorbitol-6-phosphate dehydrogenase from oral streptococci.
The activity of sorbitol-6-phosphate (S6P) dehydrogenase (S6PDH) and the sorbitol transport system were studied in strains of the oral streptococci Streptococcus gordonii, Streptococcus mitis, Streptococcus sanguis and Streptococcus mutans. Genetically transformed (to ferment sorbitol) strains and their DNA donors were included. S6PDH was purified by anion exchange chromatography and gel filtration. The purity of the enzyme was confirmed by polyacrylamide gel electrophoresis. The purified enzyme from all the strains exhibited Michaelis-Menton saturation kinetics. The Km values for nicotinamide-adenine dinucleotide (NAD) and S6P ranged between 0.03 and 0.21 mM and 0.07 and 0.20 mM respectively. The relative molecular weights of the native enzyme were 229,000 for one donor-transformant pair (S. sanguis and S. gordonii), 107,000 for the other pair (S. mitis and S. gordonii) and 129,000 for S. mutans. The molecular weights of the S6PDH subunits ranged from 26,000 to 28,000. The pH optima (greater than 8.5) and the amino acid composition (15 amino acids examined) were similar for the S6PDH from the different strains. However, the chromatographic and electrophoretic patterns as well as the Km values for NAD and S6P were the same only between the S6PDHs from the strains within each donor-transformant pair. Purified S6PDH from S. mutans also exhibited low mannitol-1-phosphate dehydrogenase activity. Sorbitol-grown decryptified cells of all the strains phosphorylated sorbitol in the presence of phosphoenolpyruvate but not in the presence of adenosine triphosphate (ATP). ATP-mediated phosphorylation of glucose was observed with the same strains when grown on glucose. No evidence for a non-phosphotransferase transport system was found for sorbitol in any of the strains.(ABSTRACT TRUNCATED AT 250 WORDS)
Svensäter
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G
Centre for Oral Health Sciences, University of Lund, Malmö, Sweden.
Edwardsson
S
S
Kalfas
S
S
D001693:Biological Transport, Active; D005947:Glucose / Q000378:metabolism; D008970:Molecular Weight; D010766:Phosphorylation; D013012:Sorbitol / Q000378:metabolism; D013291:Streptococcus / Q000201:enzymology*; D013295:Streptococcus mutans / Q000201:enzymology; D013298:Streptococcus sanguis / Q000201:enzymology; D013401:Sugar Alcohol Dehydrogenases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
D013012:Sorbitol; D013401:Sugar Alcohol Dehydrogenases; C022645:sorbitol-6-phosphate dehydrogenase; D005947:Glucose