Ribulosebisphosphate carboxylase: amino acid sequence of a peptide bearing the activator carbon dioxide

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Abstract
Ribulosebisphosphate carboxylase [from spinach] is activated by reaction of an activator CO2 to form a carbamate on the .epsilon.-amino group of a lysyl residue on the large catalytic subunit. This carbamate was converted to the methoxycarbonyl derivative by treatment of the enzyme with diazomethane, as previously reported [Lorimer, G. H. and Miziorko, H. H.]. Digestion of the methylated enzyme-14CO2 complex with trypsin yielded several radioactive peptides, which were purified by standard chromatographic procedures. Sequence analyses revealed that these peptides had the same sequence: -Gly-Gly-Leu-Asp-Phe5-Thr-Lys-Asp-Asp-Glu10-Asn-Val-Asn-Ser-Gln15-Pro-Phe. Residue 7 was 14C labeled and emerged from the sequencer as the phenylthiohydantoin derivative of N.epsilon.-(methoxycarbonyl)lysine. The acidic nature of the residues, close to the lysine bearing the activator CO2, provides a molecular explanation for the pH, and divalent metal ion dependency of the activation reaction. An entirely homologous sequence was found in the large subunit of the enzyme from Zea mays [McIntosh, L. et. al.]. The lysyl residue bearing the activator CO2 is 26 residues removed from 1 of the lysyl residues; it is identified by use of the affinity label N-bromoacetylethanolamine POH as being within the active-site domain.