Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 1993 May 15;292 ( Pt 1)(Pt 1):197-202.
doi: 10.1042/bj2920197.

Production, purification and characterization of the catalytic domain of glucoamylase from Aspergillus niger

B Stoffer  1 , T P Frandsen, P K Busk, P Schneider, I Svendsen, B Svensson
Affiliations

Production, purification and characterization of the catalytic domain of glucoamylase from Aspergillus niger

B Stoffer et al. Biochem J. .

Abstract

The catalytic domain of glucoamylases G1 and G2 from Aspergillus niger is produced in vitro in high yield by limited proteolysis using either subtilisin Novo or subtilisin Carlsberg. Purification by affinity chromatography on an acarbose-Sepharose column followed by ion-exchange chromatography on HiLoad Q-Sepharose leads to separation of a number of structurally closely related forms of domain. The cleavage occurs primarily between Val-470 and Ala-471 as indicated by C-terminal sequencing, whereas the N-terminus is intact. Subtilisin Carlsberg, in addition, produces a type of domain which is hydrolysed before Ser-444, an O-glycosylated residue. This leaves the fragment Ser-444-Val-470 disulphide-bonded to the large N-terminal part of the catalytic domain. Subtilisin Novo, in contrast, tends to yield a minor fraction of forms extending approx. 30-40 amino-acid residues beyond Val-470. The thermostability is essentially the same for the single-chain catalytic domain and the original glucoamylases G1 and G2, whereas the catalytic domain cut between Ser-443 and Ser-444 is less thermostable. For both types of domain the kinetic parameters, Km and kcat., for hydrolysis of maltose are very close to the values found for glucoamylases G1 and G2.

PubMed Disclaimer

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. J Biochem. 1976 Nov;80(5):913-22 - PubMed
    1. J Biol Chem. 1992 Sep 25;267(27):19291-8 - PubMed
    1. J Biochem. 1985 Sep;98(3):663-71 - PubMed
    1. Eur J Biochem. 1986 Feb 3;154(3):497-502 - PubMed
    1. J Bacteriol. 1987 Sep;169(9):4171-6 - PubMed

Publication types

MeSH terms

Substances