Construction of human ALAS-E expression vectors. Junction sequences of both prokaryotic (A) and eukaryotic (B) ALAS-E expression vectors were determined by sequencing. (A) wild type and mutant ALAS-E cDNA fragments were subcloned into the BamHI site of a pGEX-3X prokaryotic gene fusion vector, producing N-terminal GST fusions with Factor Xa-cleavable linkages. The papain resistant core catalytic domain of ALAS-E30 (corresponding to amino acids 125 to 587) was used in this analysis. (B) Eukaryotic expression plasmids of human wild type and mutant ALAS-E were made by inserting full-length (1.9 kb) cDNAs into the EcoRI site of pCAGGS.37 +1 indicates the cap site of ALAS-E mRNA.