Identification and molecular characterization of the first alpha -xylosidase from an archaeon.
We here report the first molecular characterization of an alpha-xylosidase (XylS) from an Archaeon. Sulfolobus solfataricus is able to grow at temperatures higher than 80 degrees C on several carbohydrates at acidic pH. The isolated xylS gene encodes a monomeric enzyme homologous to alpha-glucosidases, alpha-xylosidases, glucoamylases and sucrase-isomaltases of the glycosyl hydrolase family 31. xylS belongs to a cluster of four genes in the S. solfataricus genome, including a beta-glycosidase, an hypothetical membrane protein homologous to the major facilitator superfamily of transporters, and an open reading frame of unknown function. The alpha-xylosidase was overexpressed in Escherichia coli showing optimal activity at 90 degrees C and a half-life at this temperature of 38 h. The purified enzyme follows a retaining mechanism of substrate hydrolysis, showing high hydrolytic activity on the disaccharide isoprimeverose and catalyzing the release of xylose from xyloglucan oligosaccharides. Synergy is observed in the concerted in vitro hydrolysis of xyloglucan oligosaccharides by the alpha-xylosidase and the beta-glycosidase from S. solfataricus. The analysis of the total S. solfataricus RNA revealed that all the genes of the cluster are actively transcribed and that xylS and orf3 genes are cotranscribed.
Moracci
M
M
Institute of Protein Biochemistry and Enzymology, Consiglio Nazionale delle Ricerche (CNR), 80125 Naples, Italy. moracci@dafne.ibpe.na.cnr.it
Cobucci Ponzano
B
B
Trincone
A
A
Fusco
S
S
De Rosa
M
M
van Der Oost
J
J
Sensen
C W
CW
Charlebois
R L
RL
Rossi
M
M
D001105:Archaea / Q000201:enzymology*; D019843:Archaeal Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D005936:Glucans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000378:metabolism; D014990:Xylans; D014995:Xylosidases / Q000032:analysis* / Q000235:genetics / Q000378:metabolism
D019843:Archaeal Proteins; D005936:Glucans; D011134:Polysaccharides; D014990:Xylans; C029353:xyloglucan; D014995:Xylosidases