AUTHOR=Nurkanto Arif , Jeelani Ghulam , Yamamoto Takehiro , Hishiki Takako , Naito Yoshiko , Suematsu Makoto , Hashimoto Tetsuo , Nozaki Tomoyoshi 

TITLE=Biochemical, Metabolomic, and Genetic Analyses of Dephospho Coenzyme A Kinase Involved in Coenzyme A Biosynthesis in the Human Enteric Parasite Entamoeba histolytica

JOURNAL=Frontiers in Microbiology

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2018.02902

DOI=10.3389/fmicb.2018.02902

ISSN=1664-302X

ABSTRACT=<p>Coenzyme A (CoA) is an essential cofactor for numerous cellular reactions in all living organisms. In the protozoan parasite <italic>Entamoeba histolytica</italic>, CoA is synthesized in a pathway consisting of four enzymes with dephospho-CoA kinase (DPCK) catalyzing the last step. However, the metabolic and physiological roles of <italic>E. histolytica</italic> DPCK remain elusive. In this study, we took biochemical, reverse genetic, and metabolomic approaches to elucidate role of DPCK in <italic>E. histolytica</italic>. The <italic>E. histolytica</italic> genome encodes two DPCK isotypes (<italic>Eh</italic>DPCK1 and <italic>Eh</italic>DPCK2). Epigenetic gene silencing of <italic>Ehdpck1</italic> and <italic>Ehdpck2</italic> caused significant reduction of DPCK activity, intracellular CoA concentrations, and also led to growth retardation <italic>in vitro</italic>, suggesting importance of DPCK for CoA synthesis and proliferation. Furthermore, metabolomic analysis showed that suppression of <italic>Ehdpck</italic> gene expression also caused decrease in the level of acetyl-CoA, and metabolites involved in amino acid, glycogen, hexosamine, nucleic acid metabolisms, chitin, and polyamine biosynthesis. The kinetic properties of <italic>E. histolytica</italic> and human DPCK showed remarkable differences, e.g., the Km values of <italic>E. histolytica</italic> and human DPCK were 58–114 and 5.2 μM toward dephospho-CoA and 15–20 and 192 μM for ATP, respectively. Phylogenetic analysis also supported the uniqueness of the amebic enzyme compared to the human counterpart. These biochemical, evolutionary features, and physiological importance of <italic>Eh</italic>DPCKs indicate that <italic>Eh</italic>DPCK represents the rational target for the development of anti-amebic agents.</p>