PMID:2553397
| Citation |
Jaworski, JG, Post-Beittenmiller, MA and Ohlrogge, JB (1989) Site-directed mutagenesis of the spinach acyl carrier protein-I prosthetic group attachment site. Eur. J. Biochem. 184:603-9 |
|---|---|
| Abstract |
Site-directed mutagenesis was used to change the phosphopantetheine attachment site (Ser38) of spinach acyl carrier protein I (ACP-I) from a serine to a threonine or cysteine residue. 1. Although the native ACP-I is fully phosphopantethenylated when expressed in Escherichia coli, the TH-ACP-I and CY-ACP-I mutants were found to be completely devoid of the phosphopantetheine group. Therefore, the E. coli holoACP synthase requires serine for in vivo phosphopantetheine addition to spinach ACP-I. 2. Spinach holoACP synthase was completely inactive in vitro with either the TH-ACP-I or CY-ACP-I mutants. In addition, TH-ACP-I and CY-ACP-I were strong inhibitors of spinach holoACP synthase. 3. The mutant ACPs were weak or ineffective as inhibitors of spinach fatty acid synthesis and spinach oleoyl-ACP hydrolase. 4. Compared to holoACP-I, the mutant apoACP-I analogs had: (a) altered mobility in SDS and native gel electrophoresis, (b) altered binding to anti-(spinach ACP-I) antibodies and (c) altered isoelectric points. The combined physical, immunological and enzyme inhibition data indicate that attachment of the phosphopantheine prosthetic group alters ACP conformation. |
| Links | |
| Keywords |
Acyl Carrier Protein/analysis; Acyl Carrier Protein/genetics; Acyl Carrier Protein/isolation & purification; Cross Reactions; Electrophoresis; Escherichia coli/metabolism; Isoelectric Focusing; Mutation; Pantetheine; Phosphotransferases/antagonists & inhibitors; Plants/analysis; Protein Conformation; Transferases (Other Substituted Phosphate Groups) |
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