Cloning, expression, and characterization of the Micromonospora viridifaciens neuraminidase gene in Streptomyces lividans.

Abstract:

:We have cloned the Micromonospora viridifaciens neuraminidase (EC 3.2.1.18) gene (nedA) in Streptomyces lividans. This was accomplished by using the vector pIJ702 and BglII-BclI libraries of M. viridifaciens chromosomal inserts created in S. lividans. The libraries were screened for the expression of neuraminidase by monitoring the cleavage of the fluorogenic neuraminidase substrate 2'-(4-methylumbelliferyl)-alpha-D-N-acetyl-neuraminic acid. Positive clones (BG6, BG7, BC4, and BC8) contained the identical 2-kb BclI-BglII fragment and expressed neuraminidase efficiently and constitutively using its own promoter in the heterologous host. From the nucleotide sequence analysis, an open reading frame of 1,941 bp which encodes a polypeptide with an M(r) of 68,840 was detected. The deduced amino acid sequence has five Asp boxes, -Ser-X-Asp-X-Gly-X-Thr-Trp, showing great similarity to other bacterial and viral neuraminidases. We have also identified the catalytic domain by using truncated proteins produced in S. lividans.

journal_name

J Bacteriol

journal_title

Journal of bacteriology

authors

Sakurada K,Ohta T,Hasegawa M

doi

10.1128/jb.174.21.6896-6903.1992

keywords:

subject

Has Abstract

pub_date

1992-11-01 00:00:00

pages

6896-903

issue

21

eissn

0021-9193

issn

1098-5530

journal_volume

174

pub_type

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