Journal
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
Volume 60, Issue 36, Pages 9211-9220Publisher
AMER CHEMICAL SOC
DOI: 10.1021/jf302659z
Keywords
Dunaliella bardawil; glyceraldehyde-3-phosphate dehydrogenase (GAPDH); genome walking; regulatory element; functional diversity
Funding
- National Natural Foundation of China [31171631]
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Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a well-known proverbial protein involved in various functions in vivo. The functional diversity of GAPDH from Dunaliella bardawil (DbGAPDH) may relate to the regulatory elements lying in the promoter at the transcriptional level. Using RT-PCR and RACE reactions, gapdh cDNA was isolated, and the full-length genomic sequence was obtained by LA-PCR-based genome walking. The full-length cDNA sequence was 1645 bp containing an 1128 bp putative open reading frame (ORE), which coded a 375 amino acids-deduced polypeptide whose molecular weight was 40.27 kDa computationally. Protein conserved domain search and structural computation found that DbGAPDH consists of two structural conserved domains highly homologous in most species; multiple sequence alignment discovered two positive charge residues (Lys164 and Arg 233), which play a critical role in the protein-protein interaction between GAPDH, phosphoribulokinase (PRK), and CP12. Phylogenetic analysis demonstrated that DbGAPDH has a closer relationship with analogues from algae and higher plants than with those from other species. In silico analysis of the promoter region revealed six potential regulatory elements might be involved in four hypothesized functions characterized by chloroplast GAPDH: oxygen-, light-, pathogen-, and cold-induced regulation. These results might supply some hints for the functional diversity mechanisms of DbGAPDH, and fresh information for further research to bridge the gap between our knowledge of DNA and protein structure and our understanding of functional biology in GAPDH regulation.
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