1887

Abstract

An α-amylase-pullulanase gene from DSM 3783 was cloned in on a 7·0 kb RI fragment using a λ vector. The gene produced, from an indigenous promoter, active thermostable α-amylase-pullulanase, seemingly mostly a soluble intracellular enzyme in . Gel filtration separated the active enzyme produced into three peaks, each having both α-amylase and pullulanase activities. Immunoblotting after SDS-PAGE revealed more than ten α-amylase-pullulanase specific polypeptides; the biggest of these had an of about 165000, whereas the smallest enzymically active polypeptide had an of about 100000. Despite the marked degeneration of its constituent polypeptides, the apparent temperature optimum of the enzyme (80-85°C) was only some 5°C lower and the heat stability the same as that of the extracellular α-amylase-pullulanase produced by the native host. Oligonucleotide probes prepared according to the NH-terminal amino acid sequences of the enzyme and its satellite polypeptide (a polypeptide associated with the extracellular enzyme of the native host) hybridized to different regions of the 7·0 kb DNA insert.

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/content/journal/micro/10.1099/00221287-135-6-1755
1989-06-01
2019-11-22
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http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-135-6-1755
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