Mass spectrometric analysis of the γ subunit of Na,K-ATPase

Mass spectrometric characterisation of the two bands of the g subunit of rat renal Na,K-ATPase has been performed after SDS-PAGE purification, and development of appropriate procedures for extraction from the gel and in-gel tryptic digestion. A combination of mass determination on the two intact gamma chains, peptide mass mapping and peptide sequencing of tryptic fragments reveals that: (1) gamma1 has a mass of 7184.0 Da (carbamidomethyl cysteine) which corresponds exactly to that expected from the published sequence without the initiator methionine (Gen Bank Accession number AF129400). Thus, gamma1 is not modified, (2) gamma2 has a mass of 7353.8 Da, contains a different N-terminal sequence and is N-terminally acetylated. The seven N-terminal residues of gamma1, TELSANH, are replaced by Ac-MDRWYI/L in gamma2, but otherwise the two chains are identical.