Page 149 - Demo

SECTION I: MOLECULAR GENETIC ENGINEERING AND BIOCHEMICAL TECHNOLOGY 1533. RESULTS3.1. Sequence analysis of ncsS53The ncsS53 gene is located as a single operon with the upstream flanked to the manose-1-phosphate guanyltransferase, ncsS52, and downstream flanked to the hypothetical protein, ncsS54 [7]. The comparison of deduced amino acid sequence (333 amino acids) encoded by ncs53 with known amino acid sequences (Figure 1) showed the high similarity with the number of UDP-glucose-4-epimarase proteins involved in metabolite biosynthesis from Steptomyces: 70% identity to GalE4 [10] from S. avermitilis; 55% identity to DltE from Magnetospirillum nagnetotacticum. In addition, the deduced amino acid of ncs53showed the high homology with family of GDP-4,6-dehydratase: 49% identity to RfB3 from Leptospira interrogangs serovar lai str. 5661 and 46% to KasD from S.kasugaensis. The phylogenetic tree (Figure 2) showed that NcsS53 is the most closely related to the 4-epimerase, GalE4, from S. avermitilis. Further, sequence analysis of NcsS53 revealed two conserved motifs presenting for active sites of 4,6-dehydratase and 4-epimarase as well as; one at the vicinity of N terminal region (22VTGAAGFIGSHL33) was identified as a nucleotide-binding site [11] and the other (165S%u2026.YK171) as a NAD+ cofactor binding motif [12]. Specially, the repeated arginine residue (193RXXXXXXRXXXR204) [14] in the sequence as binding site of substrate on the enzyme indicated that ncs53 may act like 4-epimarase.