SEEKING AN ANCIENT ENZYME IN METHANOCOCCUS-JANNASCHII USING ORF, A PROGRAM BASED ON PREDICTED SECONDARY STRUCTURE COMPARISONS

We have developed a simple procedure to identify protein homologs in g enomic databases. The program, called ORF, is based on comparisons of predicted secondary structure. Protein structure is far better conserv ed than amino acid sequence, and structure-based methods have been eff ective in exploiting this fact to find homologs, even among proteins w ith scant sequence identity. ORF is a secondary structure-based method that operates solely on predictions from sequence and requires no exp erimentally determined information about the structure. The approach i s illustrated by an example: Thymidylate synthase, a highly conserved enzyme essential to thymidine biosynthesis in both prokaryotes and euk aryotes, is thought to be used by Archaea, but a corresponding gene ha s yet to be identified. Here, a candidate thymidylate synthase is iden tified as a previously unassigned open reading frame from the genome o f Methanococcus jannaschii, viz., MJ0757. Using primary structure info rmation alone, the optimally aligned sequence identity between MJ0757 and Escherichia coli thymidylate synthase is 7%, well below the thresh old of sensitivity for detection by sequence-based methods.