Identical tripeptides of the sequence X‐Pro‐Lys, where X is an unknown blocking group, were isolated from trypsin digests of bovine cardiac alkali light chain and the LC2 light chain of rabbit fast muscle. Chemical, electrophoretic and 1H‐NMR evidence characterized X as an unusual amino acid, α‐N‐trimethylalanine (Me3Ala), which was earlier reported as the N‐terminal amino acid of the A1 alkali light chain of rabbit fast muscle [Henry et al. (1982) FEBS Lett. 144, 11–15]. The narrow line width and chemical shift position (δ= 3.23 ppm) of the –N+ ‐(CH3) protons of Me3Ala made 1H‐NMR spectroscopy a convenient method to search for this residue in other light chains. A survey of many different light chains showed that this signal was present in all vertebrate striated muscle light chains of the A1‐type (LC1, ‘essential’ light chains) and LC2‐type (‘DTNB’‐light chains, ‘phosphorylatable’ light chains) but was absent from all invertebrate muscle and vertebrate smooth muscle light chains tested. It was also absent from the vertebrate fast‐muscle‐specific A2‐type (LC3) light chains. The spectral characteristics of these signals were consistent with their having arisen from the protons of an – N+ ‐(CH3)3 grouping. Since no ɛ‐trimethyllysine could be detected in acid hydrolysates of these proteins, it appears that Me3Ala is a general feature as the N‐terminal amino acid in these light chains.
1H‐NMR studies on bovine cardiac myosin subfragment 1 (S1) showed that the Me3Ala methyl proton signal was clearly visible and that the spectrum more closely resembled that of a rabbit S1 isoenzyme, S1(A1), than S1(A2), suggesting that the 40‐residue N‐terminal segment of the alkali light chain in cardiac S1 also possesses a high segmental mobility. Addition of actin caused the same gross changes to the cardiac S1 spectrum as noted earlier for rabbit S1(A1) [Prince et al. (1981) Eur. J. Biochem. 121, 213–219]. In particular, a marked reduction in the segmental mobility of the N‐terminal region of the alkali light chain was noted, consistent with a direct interaction of this area with actin.