Catalytic versatility of Bacillus pumilus. beta. -xylosidase: glycosyl transfer and hydrolysis promoted with. cap alpha. - and. beta. -D-xylosyl fluoride

Journal Article · Tue Jun 02 04:00:00 EDT 1987 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00385a009· OSTI ID:6180516

Kasumi, T; Tsumuraya, Y; Brewer, C F; Kersters-Hilderson, H; Claeyssens, M; Hehre, E J

Bacillus pumilus ..beta..-xylosidase, an enzyme considered restricted to hydrolyzing a narrow range of ..beta..-D-xylosidic substrates with inversion of configuration, was found to catalyze different stereochemical, essentially irreversible, glycosylation reactions with ..cap alpha..- and ..beta..-D-xylopyranosyl fluoride. The enzyme promoted the hydrolysis of ..beta..-D-xylopyranosyl fluoride at a high rate, V = 6.25 ..mu..mol min/sup -1/ mg/sup -1/ at 0/sup 0/C, in a reaction that obeyed Michaelis-Menten kinetics. In contrast, its action upon ..cap alpha..-D-xylopyranosyl fluoride was slow and characterized by an unusual relation between the rate of fluoride release and the substrate concentration, suggesting the possible need for two substrate molecules to be bound at the active center in order for reaction to occur. Moreover, /sup 1/H NMR spectra of a digest of ..cap alpha..-D-xylosyl fluoride showed the substrate to be specifically converted to ..cap alpha..-D-xylose by the enzyme. The observed retention of configuration is not consistent with direct hydrolysis by this inverting enzyme but is strongly indicative of the occurrence of two successive inverting reactions: xylosyl transfer from ..cap alpha..-D-xylosyl fluoride to form a ..beta..-D-xylosidic product, followed by hydrolysis of the latter to produce ..cap alpha..-D-xylose. The transient intermediate product formed enzymically from ..cap alpha..-D-xylosyl fluoride in the presence of (/sup 14/C)xylose was isolated and shown by its specific radioactivity and /sup 1/H NMR spectrum as well as by methylatino and enzymic analyses to be 4-O-..beta..-D-xylopyranosyl-D-xylopyranose containing one (/sup 14/C)xylose residue.

• Research Organization: Albert Einstein College of Medicine, Bronx, NY
• OSTI ID: 6180516
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 26:11; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

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59 BASIC BIOLOGICAL SCIENCES
62 RADIOLOGY AND NUCLEAR MEDICINE
ALDEHYDES
BACILLUS
BACTERIA
BARYONS
BIOCHEMICAL REACTION KINETICS
CARBOHYDRATES
CARBON 14 COMPOUNDS
CHEMICAL REACTIONS
CHROMATOGRAPHY
DECOMPOSITION
DEUTERIUM COMPOUNDS
ELEMENTARY PARTICLES
ENZYMATIC HYDROLYSIS
ENZYMES
FERMIONS
FLUORIDES
FLUORINE COMPOUNDS
GLYCOSYL HYDROLASES
HADRONS
HALIDES
HALOGEN COMPOUNDS
HYDROGEN COMPOUNDS
HYDROLASES
HYDROLYSIS
KINETICS
LABELLED COMPOUNDS
LYSIS
MAGNETIC RESONANCE
MICROORGANISMS
MONOSACCHARIDES
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
NUCLEONS
ORGANIC COMPOUNDS
PENTOSES
PROTONS
REACTION KINETICS
RESONANCE
SACCHARIDES
SEPARATION PROCESSES
SOLVOLYSIS
SPECTRA
STEREOCHEMISTRY
THIN-LAYER CHROMATOGRAPHY
XYLOSE