Details
| Original language | English |
|---|---|
| Pages (from-to) | 112-122 |
| Number of pages | 11 |
| Journal | Biochimica et Biophysica Acta - General Subjects |
| Volume | 1527 |
| Issue number | 3 |
| Publication status | Published - 15 Aug 2001 |
| Externally published | Yes |
Abstract
The effect of ambient pH on production and glycosylation of glucoamylase (GAM) and on the generation of a morphological mutant produced by Aspergillus niger strain B1 (a transformant containing an additional 20 copies of the homologous GAM glaA gene) was studied. We have shown that a change in the pH from 4 to 5.4 during continuous cultivation of the A. niger B1 strain instigates or accelerates the spontaneous generation of a morphological mutant (LB). This mutant strain produced approx. 50% less extracellular protein and GAM during both chemostat and batch cultivation compared to another strain with parental-type morphology (PS). The intracellular levels of GAM were also lower in the LB strain. In addition, cultivation of the original parent B1 strain in a batch-pulse bioreactor at pH 5.5 resulted in a 9-fold drop in GAM production and a 5-fold drop in extracellular protein compared to that obtained at pH 4. Glycosylation analysis of the glucoamylases purified from shake-flask cultivation showed that both principal forms of GAM secreted by the LB strain possessed enhanced galactosylation (2-fold), compared to those of the PS. Four diagnostic methods (immunostaining, mild methanolysis, mild acid hydrolysis and β-galactofuranosidase digestion) provided evidence that the majority of this galactose was of the furanoic conformation. The GAMs produced during batch-pulse cultivation at pH 5.5 similarly showed an approx. 2-fold increase in galactofuranosylation compared to pH 4. Interestingly, in both cases the increased galactofuranosylation appears primarily restricted to the O-linked glycan component. Ambient pH therefore regulates both GAM production and influences its glycosylation.
Keywords
- Aspergillus niger, Galactofuranose, Glucoamylase, pH
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biophysics
- Biochemistry, Genetics and Molecular Biology(all)
- Biochemistry
- Biochemistry, Genetics and Molecular Biology(all)
- Molecular Biology
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In: Biochimica et Biophysica Acta - General Subjects, Vol. 1527, No. 3, 15.08.2001, p. 112-122.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - The effect of pH on glucoamylase production, glycosylation and chemostat evolution of Aspergillus niger
AU - Wallis, Gregg L.F.
AU - Swift, Richard J.
AU - Atterbury, Robert
AU - Trappe, Susanne
AU - Rinas, Ursula
AU - Hemming, Frank W.
AU - Wiebe, Marilyn G.
AU - Trinci, Anthony P.J.
AU - Peberdy, John F.
PY - 2001/8/15
Y1 - 2001/8/15
N2 - The effect of ambient pH on production and glycosylation of glucoamylase (GAM) and on the generation of a morphological mutant produced by Aspergillus niger strain B1 (a transformant containing an additional 20 copies of the homologous GAM glaA gene) was studied. We have shown that a change in the pH from 4 to 5.4 during continuous cultivation of the A. niger B1 strain instigates or accelerates the spontaneous generation of a morphological mutant (LB). This mutant strain produced approx. 50% less extracellular protein and GAM during both chemostat and batch cultivation compared to another strain with parental-type morphology (PS). The intracellular levels of GAM were also lower in the LB strain. In addition, cultivation of the original parent B1 strain in a batch-pulse bioreactor at pH 5.5 resulted in a 9-fold drop in GAM production and a 5-fold drop in extracellular protein compared to that obtained at pH 4. Glycosylation analysis of the glucoamylases purified from shake-flask cultivation showed that both principal forms of GAM secreted by the LB strain possessed enhanced galactosylation (2-fold), compared to those of the PS. Four diagnostic methods (immunostaining, mild methanolysis, mild acid hydrolysis and β-galactofuranosidase digestion) provided evidence that the majority of this galactose was of the furanoic conformation. The GAMs produced during batch-pulse cultivation at pH 5.5 similarly showed an approx. 2-fold increase in galactofuranosylation compared to pH 4. Interestingly, in both cases the increased galactofuranosylation appears primarily restricted to the O-linked glycan component. Ambient pH therefore regulates both GAM production and influences its glycosylation.
AB - The effect of ambient pH on production and glycosylation of glucoamylase (GAM) and on the generation of a morphological mutant produced by Aspergillus niger strain B1 (a transformant containing an additional 20 copies of the homologous GAM glaA gene) was studied. We have shown that a change in the pH from 4 to 5.4 during continuous cultivation of the A. niger B1 strain instigates or accelerates the spontaneous generation of a morphological mutant (LB). This mutant strain produced approx. 50% less extracellular protein and GAM during both chemostat and batch cultivation compared to another strain with parental-type morphology (PS). The intracellular levels of GAM were also lower in the LB strain. In addition, cultivation of the original parent B1 strain in a batch-pulse bioreactor at pH 5.5 resulted in a 9-fold drop in GAM production and a 5-fold drop in extracellular protein compared to that obtained at pH 4. Glycosylation analysis of the glucoamylases purified from shake-flask cultivation showed that both principal forms of GAM secreted by the LB strain possessed enhanced galactosylation (2-fold), compared to those of the PS. Four diagnostic methods (immunostaining, mild methanolysis, mild acid hydrolysis and β-galactofuranosidase digestion) provided evidence that the majority of this galactose was of the furanoic conformation. The GAMs produced during batch-pulse cultivation at pH 5.5 similarly showed an approx. 2-fold increase in galactofuranosylation compared to pH 4. Interestingly, in both cases the increased galactofuranosylation appears primarily restricted to the O-linked glycan component. Ambient pH therefore regulates both GAM production and influences its glycosylation.
KW - Aspergillus niger
KW - Galactofuranose
KW - Glucoamylase
KW - pH
UR - http://www.scopus.com/inward/record.url?scp=0035882465&partnerID=8YFLogxK
U2 - 10.1016/S0304-4165(01)00145-3
DO - 10.1016/S0304-4165(01)00145-3
M3 - Article
C2 - 11479027
AN - SCOPUS:0035882465
VL - 1527
SP - 112
EP - 122
JO - Biochimica et Biophysica Acta - General Subjects
JF - Biochimica et Biophysica Acta - General Subjects
SN - 0304-4165
IS - 3
ER -