ATP-dependent Saccharomyces cerevisiae phospho enol pyruvate carboxykinase: isolation and sequence of a peptide containing a highly reactive cysteine.
Saccharomyces cerevisiae phospho enol pyruvate carboxykinase (EC 4.1.1.49), inactivated by N-(iodoacetyl)-N'-(5-sulfo-1-naphthyl)ethylenediamine, incorporated 0.95 mol of the fluorescent moiety per mol of enzyme subunit. Reagent incorporation was completely protected by the presence of ADP plus MnCl2. The labeled protein was digested with trypsin after carboxymethylation. Two labeled peptides were isolated by reverse-phase high-performance liquid chromatography and were sequenced by gas-phase automatic Edman degradation. Both peptides contained overlapping amino acid sequences from Asn-358 to Lys-375, thus identifying Cys-364 as the reactive amino acid residue. The position of the target amino acid residue is immediately preceding a putative phosphoryl-binding sequence proposed for some nucleotide-binding proteins.
Alvear
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M
Departamento de Ciencias Químicas, Facultad de Ingeniería y Administración, Universidad de la Frontera, Temuco, Chile.
Encinas
M V
MV
Kemp
R G
RG
Latshaw
S P
SP
Cardemil
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E
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D003545:Cysteine; D004926:Escherichia coli / Q000201:enzymology; D005456:Fluorescent Dyes; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009282:Naphthalenesulfonates; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010729:Phosphoenolpyruvate Carboxykinase (GTP) / Q000737:chemistry* / Q000378:metabolism; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D012689:Sequence Homology, Nucleic Acid; D013050:Spectrometry, Fluorescence; D014346:Trypanosoma brucei brucei / Q000201:enzymology
D005456:Fluorescent Dyes; D046911:Macromolecular Substances; D009282:Naphthalenesulfonates; D010446:Peptide Fragments; C006421:1,5-I-AEDANS; D000255:Adenosine Triphosphate; D010729:Phosphoenolpyruvate Carboxykinase (GTP); D003545:Cysteine