Abstract
A cDNA expression library of Trichoderma reesei RutC-30 was constructed in the yeast Saccharomyces cerevisiae. Two genes, abfI and bxII, were isolated by screening the yeast library for extracellular α-L- arabinofuranosidase activity with the substrate p-nitrophenyl-α-L- arabinofuranoside. The genes abfI and bxII encode 500 and 758 amino acids, respectively, including the signal sequences. The deduced amino acid sequence of ABFI displays high-level similarity to the α-L-arabinofuranosidase B of Aspergillus niger, and the two can form a new family of glycosyl hydrolases. The deduced amino acid sequence of BXLI shows similarities to the β- glucosidases grouped in family 3. The yeast-produced enzymes were tested for enzymatic activities against different substrates. ABFI released L-arabinose from p-nitrophenyl-α-L-arabinofuranoside and arahinoxylans and showed some β-xylosidase activity toward p-nitrophenyl-β-D-xylopyranoside. BXLI did not release L-arabinose from arabinoxylan. It showed α-L-arabinofuranosidase, α-L-arabinopyranosidase, and β-xylosidase activities against p- nitrophenyl-α-L-arahinofuranosidase, p-nitrophenyl-α-arabinopyranoside, and p-nitrophenyl-β-D-xylopyranoside, respectively, with the last activity being the highest. It was also able to hydrolyze xyhobiose and slowly release xylose from polymeric xylan. ABFI and BXLI correspond to a previously purified α-L-arabinofuranosidase and a β-xylosidase from T. reesei, respectively, as confirmed by partial amino acid sequencing of the Trichoderma-produced enzymes. Both enzymes produced in yeasts displayed hydrolytic properties similar to those of the corresponding enzymes purified from T. reesei.
| Original language | English |
|---|---|
| Pages (from-to) | 3840-3846 |
| Number of pages | 7 |
| Journal | Applied and Environmental Microbiology |
| Volume | 62 |
| Issue number | 10 |
| Publication status | Published - 1 Oct 1996 |
| MoE publication type | A1 Journal article-refereed |
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Keywords
- 4 nitrophenol
- alpha arabinofuranosidase
- pyranoside
- Xylan
- xylan 1,4 beta xylosidase
- Xylose
- Amino Acid Sequence
- Aspergillus niger
- Hypocrea jecorina
- Saccharomyces cerevisiae
- Trichoderma
Cite this
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Cloning of genes encoding α-L-arabinofuranosidase and β-xylosidase from Trichoderma reesei by expression in Saccharomyces cerevisiae. / Margolles-Clark, Emilio; Tenkanen, Maija; Nakari-Setälä, Tiina; Penttilä, Merja.
In: Applied and Environmental Microbiology, Vol. 62, No. 10, 01.10.1996, p. 3840-3846.Research output: Contribution to journal › Article › Scientific › peer-review
TY - JOUR
T1 - Cloning of genes encoding α-L-arabinofuranosidase and β-xylosidase from Trichoderma reesei by expression in Saccharomyces cerevisiae
AU - Margolles-Clark, Emilio
AU - Tenkanen, Maija
AU - Nakari-Setälä, Tiina
AU - Penttilä, Merja
PY - 1996/10/1
Y1 - 1996/10/1
N2 - A cDNA expression library of Trichoderma reesei RutC-30 was constructed in the yeast Saccharomyces cerevisiae. Two genes, abfI and bxII, were isolated by screening the yeast library for extracellular α-L- arabinofuranosidase activity with the substrate p-nitrophenyl-α-L- arabinofuranoside. The genes abfI and bxII encode 500 and 758 amino acids, respectively, including the signal sequences. The deduced amino acid sequence of ABFI displays high-level similarity to the α-L-arabinofuranosidase B of Aspergillus niger, and the two can form a new family of glycosyl hydrolases. The deduced amino acid sequence of BXLI shows similarities to the β- glucosidases grouped in family 3. The yeast-produced enzymes were tested for enzymatic activities against different substrates. ABFI released L-arabinose from p-nitrophenyl-α-L-arabinofuranoside and arahinoxylans and showed some β-xylosidase activity toward p-nitrophenyl-β-D-xylopyranoside. BXLI did not release L-arabinose from arabinoxylan. It showed α-L-arabinofuranosidase, α-L-arabinopyranosidase, and β-xylosidase activities against p- nitrophenyl-α-L-arahinofuranosidase, p-nitrophenyl-α-arabinopyranoside, and p-nitrophenyl-β-D-xylopyranoside, respectively, with the last activity being the highest. It was also able to hydrolyze xyhobiose and slowly release xylose from polymeric xylan. ABFI and BXLI correspond to a previously purified α-L-arabinofuranosidase and a β-xylosidase from T. reesei, respectively, as confirmed by partial amino acid sequencing of the Trichoderma-produced enzymes. Both enzymes produced in yeasts displayed hydrolytic properties similar to those of the corresponding enzymes purified from T. reesei.
AB - A cDNA expression library of Trichoderma reesei RutC-30 was constructed in the yeast Saccharomyces cerevisiae. Two genes, abfI and bxII, were isolated by screening the yeast library for extracellular α-L- arabinofuranosidase activity with the substrate p-nitrophenyl-α-L- arabinofuranoside. The genes abfI and bxII encode 500 and 758 amino acids, respectively, including the signal sequences. The deduced amino acid sequence of ABFI displays high-level similarity to the α-L-arabinofuranosidase B of Aspergillus niger, and the two can form a new family of glycosyl hydrolases. The deduced amino acid sequence of BXLI shows similarities to the β- glucosidases grouped in family 3. The yeast-produced enzymes were tested for enzymatic activities against different substrates. ABFI released L-arabinose from p-nitrophenyl-α-L-arabinofuranoside and arahinoxylans and showed some β-xylosidase activity toward p-nitrophenyl-β-D-xylopyranoside. BXLI did not release L-arabinose from arabinoxylan. It showed α-L-arabinofuranosidase, α-L-arabinopyranosidase, and β-xylosidase activities against p- nitrophenyl-α-L-arahinofuranosidase, p-nitrophenyl-α-arabinopyranoside, and p-nitrophenyl-β-D-xylopyranoside, respectively, with the last activity being the highest. It was also able to hydrolyze xyhobiose and slowly release xylose from polymeric xylan. ABFI and BXLI correspond to a previously purified α-L-arabinofuranosidase and a β-xylosidase from T. reesei, respectively, as confirmed by partial amino acid sequencing of the Trichoderma-produced enzymes. Both enzymes produced in yeasts displayed hydrolytic properties similar to those of the corresponding enzymes purified from T. reesei.
KW - 4 nitrophenol
KW - alpha arabinofuranosidase
KW - pyranoside
KW - Xylan
KW - xylan 1,4 beta xylosidase
KW - Xylose
KW - Amino Acid Sequence
KW - Aspergillus niger
KW - Hypocrea jecorina
KW - Saccharomyces cerevisiae
KW - Trichoderma
UR - http://www.scopus.com/inward/record.url?scp=0029739169&partnerID=8YFLogxK
M3 - Article
VL - 62
SP - 3840
EP - 3846
JO - Applied and Environmental Microbiology
JF - Applied and Environmental Microbiology
SN - 0099-2240
IS - 10
ER -