Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.3.1 (citrate synthase)
 4,488 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The chromatographic separation of amino acids for compositional analysis of peptides and proteins is commonly performed by reverse-phase high-performance liquid chromatography of amino acid residues that have been derivatized with phenylisothiocyanate. The present report describes an extension of this method, which employs thermospray liquid chromatography/mass spectrometry to confirm the identification of the resulting phenylthiocarbamyl (PTC) amino acids. A standard HPLC separation method has been adapted for use with the thermospray technique, and on-column mass spectra of standard synthetic PTC-amino acids have been acquired. These spectra show characteristic fragmentation patterns not seen in the corresponding cyclic phenylthiohydantoin amino acid derivatives. The LC/MS method has been tested on hydrolysates of bovine serum albumin, porcine insulin, and human placental collagen. In each case, the mass spectra of components eluting with the same retention times as the standard PTC-amino acids are similar to those observed in the standard amino acid mixture. Other components display mass spectra that can be interpreted in terms of known in vivo or in vitro modifications to amino acid side chains in these proteins. The LC/MS method has assisted in the identification of by-products of the derivatization reaction. It has also been applied to a study in which an enzyme, citrate synthase, isolated from porcine heart, was compared to the protein expressed by a recombinant porcine citrate synthase gene in Escherichia coli. The data showed that the recombinant protein lacks a modified residue, trimethyllysine, which is present in the enzyme expressed in mammalian tissues.
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PMID:Identification of phenylthiocarbamyl amino acids for compositional analysis by thermospray liquid chromatography/mass spectrometry. 266 8

Detailed evidence for the amino acid sequence of allosteric citrate synthase from Escherichia coli is presented. The evidence confirms all but 11 of the residues inferred from the sequence of the gene as reported previously [Ner, S. S., Bhayana, V., Bell, A. W., Giles, I. G., Duckworth, H. W., & Bloxham, D. P. (1983) Biochemistry 22, 5243]; no information has been obtained about 10 of these (residues 101-108 and 217-218), and we find aspartic acid rather than asparagine at position 10. Substantial regions of sequence homology are noted between the E. coli enzyme and citrate synthase from pig heart, especially near residues thought to be involved in the active site. Deletions or insertions must be assumed in a number of places in order to maximize homology. Either of two lysines, at positions 355 and 356, could be formally homologous to the trimethyllysine of pig heart enzyme, but neither of these is methylated. It appears that E. coli and pig heart citrate synthases are formed of basically similar subunits but that considerable differences exist, which must explain why the E. coli enzyme is hexameric and allosterically inhibited by NADH, while the pig heart enzyme is dimeric and insensitive to that nucleotide.
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PMID:Amino acid sequence of Escherichia coli citrate synthase. 638 May 76

The sequence of 437 amino acid residues of porcine heart citrate synthase [citrate oxaloacetate-lyase (pro-3S-CH2COO leads to acetyl-CoA), EC 4. 1. 3. 7] has been determined by the alignment of fragments generated by cleavage with cyanogen bromide and with trypsin. Isolation of the peptides was facilitated by recent developments in the high-performance liquid chromatography of peptide mixtures. The alignment of these peptides was consistent with that previously deduced from fragments derived by restricted cleavage of citrate synthase by limited proteolysis and cleavage of aspartyl-prolyl bonds and asparaginyl-glycyl bonds. The enzyme contains a modified amino acid, trimethyllysine, at residue 368, showing that the enzyme is subjected to post-translational modification.
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PMID:Primary structure of porcine heart citrate synthase. 679 32

The detailed proof of the 437-residue amino acid sequence (Mr 48,969) of porcine heart citrate synthase (EC 4.13.7) is described. The S-carboxymethylated protein has been cleaved at methionine (cyanogen bromide) and arginine (trypsin digest of citraconylated enzyme) residues to yield 14 and 17 major peptides, respectively. Peptides were initially fractionated by gel filtration, and those useful for sequence analysis were purified by high-performance liquid chromatography. Sequence analyses were performed on these primary peptides and on subpeptides generated by cleavage with the bromine adduct of 2-[(2-nitrophenyl)sulfenyl]-3-methylindole, Staphylococcus aureus V8 protease, trypsin, chymotrypsin, or acid. The overall sequence was confirmed by analyzing products of cleavage by hydroxylamine, acid, and subtilisin. A novel feature of the sequence is the identification of trimethyllysine at residue 368.
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PMID:Complete amino acid sequence of porcine heart citrate synthase. 709 27