The occurrence of a main amino acid at the start of proteins from various vertebrate species is reported. Computer analyses of protein-coding sequences showed that alanine occurs in one out of five cases as the second amino acid in proteins from seven mammals (including man), one amphibian, one bird and two fishes. We also show that the alanine codon GCG occurs >3-fold more often as second codon than in general, i.e. GCG was overrepresented. Options to explain the abundance of alanine as second amino acid may already exist, but the overrepresentation of GCG was harder to explain because GCG was classically a rare codon. However, based on similarities with the published translation-initiation enhancer sequence CCGGCGG, which has complementarity with 18S ribosomal RNA, a theoretical role of GCG in translation initiation is suggested. Namely, GCG is proposed to be part of a sequence with potential for base-pairing with 18S ribosomal RNA. (C) 2008 Elsevier Inc. All rights reserved.