Secretion of the Alpha-galactosidase from Cyamopsis Tetragonoloba (guar) by Bacillus Subtilis

Overview

Journal Appl Environ Microbiol

Specialties Environmental Health
Microbiology

Date 1990 May 1

PMID 2160224

Citations 8

Authors

N Overbeeke

G H Termorshuizen

M L Giuseppin

D R Underwood

C T Verrips

Affiliations

Soon will be listed here.

Abstract

A fusion of DNA sequences encoding the SPO2 promoter, the alpha-amylase signal sequence from Bacillus amyloliquefaciens, and the mature part of the alpha-galactosidase from Cyamopsis tetragonoloba (guar) was constructed on a Bacillus subtilis multicopy vector. Bacillus cells of the protease-deficient strain DB104 harboring this vector produced and secreted the plant enzyme alpha-galactosidase up to levels of 1,700 U/liter. A growth medium suppressing the residual proteolytic activity of strain DB104 was used to reach these levels in a fermentor. Purification of the secreted product followed by NH2-terminal amino acid sequencing showed that the alpha-amylase signal sequence had been processed correctly. The molecular mass of the product estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was slightly lower than that of the plant purified enzyme, which is most likely due to glycosylation of the latter. The alpha-galactosidase product was active both on the artificial substrate para-nitrophenyl-alpha-D-galactopyranoside and on the galactomannan substrate, guar gum. The activity of this Bacillus sp.-produced enzyme was similar to that of the glycosylated enzyme purified from guar seeds, indicating that glycosylation has no essential function for enzyme activity.

Citing Articles

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Glucoamylase gene fusions alleviate limitations for protein production in Aspergillus awamori at the transcriptional and (post) translational levels.

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Analysis of heterologous protein production in defined recombinant Aspergillus awamori strains.

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Protein secretion in Bacillus species.

Simonen M, PALVA I Microbiol Rev. 1993; 57(1):109-37.

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Comparative study on the production of guar alpha-galactosidase by Saccharomyces cerevisiae SU50B and Hansenula polymorpha 8/2 in continuous cultures.

Giuseppin M, Almkerk J, Heistek J, Verrips C Appl Environ Microbiol. 1993; 59(1):52-9.

PMID: 8382463 PMC: 202054. DOI: 10.1128/aem.59.1.52-59.1993.

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