Metabolism of sucrose and its five linkage-isomeric alpha-D-glucosyl-D-fructoses by Klebsiella pneumoniae. Participation and properties of sucrose-6-phosphate hydrolase and phospho-alpha-glucosidase.
Klebsiella pneumoniae is presently unique among bacterial species in its ability to metabolize not only sucrose but also its five linkage-isomeric alpha-d-glucosyl-d-fructoses: trehalulose, turanose, maltulose, leucrose, and palatinose. Growth on the isomeric compounds induced a protein of molecular mass approximately 50 kDa that was not present in sucrose-grown cells and which we have identified as an NAD(+) and metal ion-dependent 6-phospho-alpha-glucosidase (AglB). The aglB gene has been cloned and sequenced, and AglB (M(r) = 49,256) has been purified from a high expression system using the chromogenic p-nitrophenyl alpha-glucopyranoside 6-phosphate as substrate. Phospho-alpha-glucosidase catalyzed the hydrolysis of a wide variety of 6-phospho-alpha-glucosides including maltose-6'-phosphate, maltitol-6-phosphate, isomaltose-6'-phosphate, and all five 6'-phosphorylated isomers of sucrose (K(m) approximately 1-5 mm) yet did not hydrolyze sucrose-6-phosphate. By contrast, purified sucrose-6-phosphate hydrolase (M(r) approximately 53,000) hydrolyzed only sucrose-6-phosphate (K(m) approximately 80 microm). Differences in molecular shape and lipophilicity potential between sucrose and its isomers may be important determinants for substrate discrimination by the two phosphoglucosyl hydrolases. Phospho-alpha-glucosidase and sucrose-6-phosphate hydrolase exhibit no significant homology, and by sequence-based alignment, the two enzymes are assigned to Families 4 and 32, respectively, of the glycosyl hydrolase superfamily. The phospho-alpha-glucosidase gene (aglB) lies adjacent to a second gene (aglA), which encodes an EII(CB) component of the phosphoenolpyruvate-dependent sugar:phosphotransferase system. We suggest that the products of the two genes facilitate the phosphorylative translocation and subsequent hydrolysis of the five alpha-d-glucosyl-d-fructoses by K. pneumoniae.
Thompson
J
J
Microbial Biochemistry and Genetics Unit, Oral Infection and Immunity Branch, NIDCR, National Institutes of Health, Bethesda, Maryland 20892, USA. jthompson@dir.nidcr.noh.gov
Robrish
S A
SA
Immel
S
S
Lichtenthaler
F W
FW
Hall
B G
BG
Pikis
A
A
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000302:isolation & purification; D001483:Base Sequence; D001692:Biological Transport; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis; D004926:Escherichia coli / Q000201:enzymology; D005632:Fructose / Q000737:chemistry; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006868:Hydrolysis; D007711:Klebsiella pneumoniae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D008670:Metals / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013395:Sucrose / Q000378:metabolism*; D000520:alpha-Glucosidases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D043324:beta-Fructofuranosidase
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008670:Metals; D020033:Protein Isoforms; D005632:Fructose; D013395:Sucrose; D006026:Glycoside Hydrolases; C049150:maltose-6'-phosphate glucosidase; D000520:alpha-Glucosidases; D043324:beta-Fructofuranosidase