| Summary: | 碩士 === 國立臺灣大學 === 農業化學系 === 82 === Streptomyces rimosus TM-55 was treated with 3% ethyl methane
sulfonate for 60 minutes. Two mutants with high amylase
activity were selected from 1,283 2-deoxyglucose depressed
mutants. In submerged fermentation, we found that
2-deoxyglucose and lactose acted as inducers for amylase
production. Purification of S. rimosus TM-55 α-amylase was
achived by ammonium sulfate precipitation, DEAE-Sephacel ion
exchange column and Sephadex G-100 gel filtration column. The
recovery of purifi- ed α-amylase was 11.07% and purification
factor is 23.64 fold. The enzyme has a molecular mass 65 kDa.
Furthermore, we cloned the S. rimosus TM-55 α-amylase gene
(amy). Its open reading frame has 1776 bp and encodes a protein
containing 592 amino acid residues with molecular mass of 65,7
kDa. This protein containes four highly conserved domains which
connect to starch binding sites and catalytic sites.
Transformation of high copy number recombinant plasmids
containing amylase gene into S. rimosus TM-55 was followed by
submerged fermentation. We found that the amylase activity in
transformants is 1.7-2.0 fold higher than that of wild type;
and the oxytetracycline titer in the fermentation broth of the
transformants is also 2 fold higher than that of the wild type,
S. rimosus TM-55.
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