STRUCTURE OF THE GENE ENCODING LAMINARIPENTAOSE-PRODUCING BETA-1,3-GLUCANASE (LPHASE) OF STREPTOMYCES-MATENSIS DIC-108

Streptomyces matensis DIC-108 secretes a laminaripentaose-producing be ta-1,3-glucanase (LPHase) into the culture medium. LPHase was partiall y purified and its N-terminal amino acid sequence was determined. The gene (Iph) for LPHase was cloned using a colony hybridization techniqu e and a synthetic oligonucleotide probe designed from the N-terminal a mino acid sequence, and the nucleotide sequence of Eph was determined. Iph has an open reading frame of 1203 bp that encode a polypeptide of 401 amino acids including a putative 35-amino acid signal sequence. T he mature portion of the polypeptide showed significant sequence simil arity with the catalytic domains of beta-1,3-glucanases of Oerskovia x anthineolytica and Arthrobacter sp. strain YCWD3. Since Iph was not ex pressed in Escherichia coli, the coding region of Iph was amplified by PCR and ligated into the expression vector pKK233-2. E. coil JM109 ce lls carrying the resultant plasmid pKK-lph produced beta-1,3-glucanase of a size identical to that of LPHase produced by S. matensis DIC-108 , and the enzyme exclusively liberated laminaripentaose from insoluble beta-l,3-glucan.