Glutamine as a Precursor to N-Terminal Pyrrolid-2-one- 5-carboxylic Acid in Mouse Immunoglobulin A-Type Light Chains AMINO ACID-SEQUENCE VARIABILITY AT THE N-TERMINAL EXTRA PIECE OF A-TYPE LIGHT-CHAIN PRECURSORS By YIGAL BURSTEIN and ISRAEL SCHECHTER

The mRNA molecules coding for three mouse immunoglobulin A-type light (L) chains (MOPC-104E Al, RPC-20 Al, MOPC-315 A2) programme the cell-free synthesis of precursors larger than the mature proteins. Radioactive amino acid-sequence analyses of each of the three precursors labelled with [3H]alanine, [3H]serine, [3H]glutamine, [3H]glutamic acid and [3H]threonine showed that an extra piece, at least 18 residues long, is linked to the N-terminus ofthe mature L-chains. The N-terminal extra-peptide segment may be 19 residues long, since analyses of precursors labelled with [35S]methionine indicated an additional N-terminal methionine residue which was recovered in low yields. Presumably this is the initiator methionine, which is known to be short lived in eukaryotes. The mature forms of MOPC-104E, RPC-20 and MOPC-315 A L-chains are blocked at the N-termini by pyrrolid-2-one-5-carboxylic acid (pyroglutamic acid). Sequence analyses of precursors labelled with [3H]glutamine and [3H]glutamic acid showed incorporation only of glutamine in a position that matches with the position of pyrrolid-2-one-5-carboxylic acid in the mature forms of all three precursors, and incorporation ofglutamic acid in other positions. The data showed the absence ofglutamine-glutamic acid interconversion, since the radioactive peaks obtained from either 3Hlabelled amino acid were discrete, and free from cross-contamination. These results prove that glutamine is the precursor amino acid of pyrrolid-2-one-5-carboxylic acid at the N-termini of the mature MOPC-104E Al, RPC-20 Al and MOPC-315 A2 L-chains. Thus the formation of pyrrolid-2-one-5-carboxylic acid by cyclization of glutamine is a post-translational event which occurs after, or concomitant with, cleavage of the extra piece from the precursor to yield the mature L-chain. The variable (V) regions (110 amino acid residues) ofmouse A L-chains are quite similar: when compared with that of MOPC104E Al chain, the V-region of RPC-20 A1 chain differs in one residue, and the V-region of MOPC-315 A2 chain differs in 11 residues. The partial sequence data show that the N-terminal extra pieces of the two Al L-chain precursors have, so far, identical partial sequences; the extra piece of the A2 L-chain precursor differs from these in at least three out of 19 positions.