Details
| Original language | English |
|---|---|
| Pages (from-to) | 1951-1958 |
| Number of pages | 8 |
| Journal | CHEMBIOCHEM |
| Volume | 7 |
| Issue number | 12 |
| Publication status | Published - 1 Dec 2006 |
| Externally published | Yes |
Abstract
Norsolorinic acid synthase (NSAS) is a type I iterative polyketide synthase that occurs in the filamentous fungus Aspergillus parasiticus. PCR was used to clone fragments of NSAS corresponding to the acyl carrier protein (ACP), acyl transferase (AT) and β-ketoacyl-ACP synthase (KS) catalytic domains. Expression of these gene fragments in Escherichia coli led to the production of soluble ACP and AT proteins. Coexpression of ACP with E. coli holo-ACP synthase (ACPS) let to production of NSAS holo-ACP, which could also be formed in vitro by using Streptomyces coelicolor ACPS. Analysis by mass spectrometry showed that, as with other type I carrier proteins, self-malonylation is not observed in the presence of malonyl CoA alone. However, the NSAS holo-ACP serves as substrate for S. coelicotor MCAT, S. coelicolor actinorhodin holo-ACP and NSAS AT domain-catalysed malonate transfer from malonyl CoA. The AT domain could transfer malonate from malonyl CoA to NSAS holo-ACP, but not hexanoate or acetate from either the cognate CoA or FAS ACP species to NSAS holo-ACP. The NSAS holo-ACP was also active in actinorhodin minimal PKS assays, but only in the presence of exogenous malonyl transferases.
Keywords
- Actinorhodin, Enzymes, Norsolorinic acid, Polyketide synthase, Proteins
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biochemistry
- Biochemistry, Genetics and Molecular Biology(all)
- Molecular Medicine
- Biochemistry, Genetics and Molecular Biology(all)
- Molecular Biology
- Chemistry(all)
- Organic Chemistry
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In: CHEMBIOCHEM, Vol. 7, No. 12, 01.12.2006, p. 1951-1958.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Catalytic relationships between type I and type II iterative polyketide synthases
T2 - The Aspergillus parasiticus norsolorinic acid synthase
AU - Ma, Yue
AU - Smith, Leah H.
AU - Cox, Russell J.
AU - Beltran-Alvarez, Pedro
AU - Arthur, Christopher J.
AU - Simpson, Thomas J.
PY - 2006/12/1
Y1 - 2006/12/1
N2 - Norsolorinic acid synthase (NSAS) is a type I iterative polyketide synthase that occurs in the filamentous fungus Aspergillus parasiticus. PCR was used to clone fragments of NSAS corresponding to the acyl carrier protein (ACP), acyl transferase (AT) and β-ketoacyl-ACP synthase (KS) catalytic domains. Expression of these gene fragments in Escherichia coli led to the production of soluble ACP and AT proteins. Coexpression of ACP with E. coli holo-ACP synthase (ACPS) let to production of NSAS holo-ACP, which could also be formed in vitro by using Streptomyces coelicolor ACPS. Analysis by mass spectrometry showed that, as with other type I carrier proteins, self-malonylation is not observed in the presence of malonyl CoA alone. However, the NSAS holo-ACP serves as substrate for S. coelicotor MCAT, S. coelicolor actinorhodin holo-ACP and NSAS AT domain-catalysed malonate transfer from malonyl CoA. The AT domain could transfer malonate from malonyl CoA to NSAS holo-ACP, but not hexanoate or acetate from either the cognate CoA or FAS ACP species to NSAS holo-ACP. The NSAS holo-ACP was also active in actinorhodin minimal PKS assays, but only in the presence of exogenous malonyl transferases.
AB - Norsolorinic acid synthase (NSAS) is a type I iterative polyketide synthase that occurs in the filamentous fungus Aspergillus parasiticus. PCR was used to clone fragments of NSAS corresponding to the acyl carrier protein (ACP), acyl transferase (AT) and β-ketoacyl-ACP synthase (KS) catalytic domains. Expression of these gene fragments in Escherichia coli led to the production of soluble ACP and AT proteins. Coexpression of ACP with E. coli holo-ACP synthase (ACPS) let to production of NSAS holo-ACP, which could also be formed in vitro by using Streptomyces coelicolor ACPS. Analysis by mass spectrometry showed that, as with other type I carrier proteins, self-malonylation is not observed in the presence of malonyl CoA alone. However, the NSAS holo-ACP serves as substrate for S. coelicotor MCAT, S. coelicolor actinorhodin holo-ACP and NSAS AT domain-catalysed malonate transfer from malonyl CoA. The AT domain could transfer malonate from malonyl CoA to NSAS holo-ACP, but not hexanoate or acetate from either the cognate CoA or FAS ACP species to NSAS holo-ACP. The NSAS holo-ACP was also active in actinorhodin minimal PKS assays, but only in the presence of exogenous malonyl transferases.
KW - Actinorhodin
KW - Enzymes
KW - Norsolorinic acid
KW - Polyketide synthase
KW - Proteins
UR - http://www.scopus.com/inward/record.url?scp=33845449100&partnerID=8YFLogxK
U2 - 10.1002/cbic.200600341
DO - 10.1002/cbic.200600341
M3 - Article
C2 - 17086560
AN - SCOPUS:33845449100
VL - 7
SP - 1951
EP - 1958
JO - CHEMBIOCHEM
JF - CHEMBIOCHEM
SN - 1439-4227
IS - 12
ER -