Document Detail

Molecular cloning and functional analysis of (R)-3-hydroxyacyl-acyl carrier protein:coenzyme A transacylase from Pseudomonas mendocina LZ.
MedLine Citation:
PMID:  16213672     Owner:  NLM     Status:  MEDLINE    
An inactive (R)-3-hydroxyacyl-acyl carrier protein:coenzyme A transacylase (PhaG(Pm)) was cloned from a newly isolated Proteobacteria Pseudomonas mendocina LZ. It is the first characterized native inactive PhaG protein. Sequence analysis indicated that there were only two sites where the amino acid sequence differed between this inactive protein and the functional PhaG(Pp) from P. putida. The differences were located at position 78 and in the region 109-113 in the amino acid sequence. Mutagenesis was carried out to investigate these two sites. A recombinant strain harboring a S78C PhaG(Pp) mutant accumulated polyhydroxyalkanoates (PHA) at 11.9% of the cellular dry weight, as compared to the 21.6% PHA produced by the recombinant harboring the wild-type PhaG(Pp). On the other hand, the changes in the amino acid region 109-113 of PhaG(Pp) to its corresponding region of PhaG(Pm) resulted in negligible PHA accumulation. This demonstrated that region 109-113 in PhaG is relatively important for transacylase activity, while position 78 just plays a supporting role for the enzyme. Furthermore, 3-D structural models of PhaG(Pp) and PhaG(Pm) developed by computational prediction revealed that the variation in amino acids at 109-113 leads to the destruction of the PhaG catalytic center, resulting in the loss of enzyme activity.
Leo Zhong Zheng; Zhi Li; Hong-Lei Tian; Ming Li; Guo-Qiang Chen
Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2005-09-22
Journal Detail:
Title:  FEMS microbiology letters     Volume:  252     ISSN:  0378-1097     ISO Abbreviation:  FEMS Microbiol. Lett.     Publication Date:  2005 Nov 
Date Detail:
Created Date:  2005-10-31     Completed Date:  2006-03-16     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  7705721     Medline TA:  FEMS Microbiol Lett     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  299-307     Citation Subset:  IM    
MOE laboratory of Protein Science, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing, China.
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MeSH Terms
Acyltransferases / chemistry*,  genetics*
Amino Acid Sequence
Amino Acid Substitution
Bacterial Proteins / genetics
Catalytic Domain
Cloning, Molecular*
DNA, Bacterial / chemistry,  genetics
Genes, Bacterial
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutation, Missense
Protein Conformation
Pseudomonas mendocina / enzymology*,  genetics
Sequence Analysis, DNA
Sequence Homology, Amino Acid
Reg. No./Substance:
0/Bacterial Proteins; 0/DNA, Bacterial; EC 2.3.-/Acyltransferases; EC 2.3.1.-/3-hydroxyacyl-CoA-acyl carrier protein transferase

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