The role of acetate fermentation in Entamoeba histolytica growth and infection

醋酸发酵在溶组织内阿米巴生长和感染中的作用

• 批准号: 9900819
• 负责人: CHERYL Jean INGRAM-SMITH
• 金额: $ 22.09万
• 依托单位: CLEMSON UNIVERSITY
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至 2022-07-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9900819
• 关键词: Acetate-CoA Ligase; Acetates; Acetyl Coenzyme A; Actins; Active Sites; Anaerobic Bacteria; Bacteria; Basidiomycota; Biochemistry; Bioterrorism; Blastocystis; Category B pathogen; Cause of Death; Cells; Centers of Research Excellence; Cessation of life; Citric Acid Cycle; Coenzyme A; Colon; Cryptosporidium; Cyst; Developing Countries; Dose; Energy Metabolism; Energy Metabolism Pathway; Entamoeba; Entamoeba histolytica; Enzymatic Biochemistry; Enzymes; Evolution; Fermentation; Food Supply; Generations; Giardia; Glucose; Glycolysis; Goals; Growth; Heat-Shock Response; Human; Infection; Investigation; Life Cycle Stages; Metabolic; Metabolic Pathway; Microbe; Mitochondria; Morbidity - disease rate; National Institute of Allergy and Infectious Disease; Organism; Orthophosphate Dikinase Pyruvate; Oxidative Phosphorylation; Parasites; Parasitic Diseases; Pathogenicity; Pathway interactions; Phosphorylation; Phosphotransferases; Physiological; Physiology; Plasmodium; Play; Production; Property; Pyruvate; Pyruvate Kinase; Pyruvate synthase; RNA Interference; Reaction; Recombinants; Research; Research Personnel; Role; Source; Structure; Succinate-CoA Ligases; Water Supply; Work; acetyl phosphate; axenic culture; cell motility; combat; enzyme structure; fungus; in vivo; innovation; insight; interest; mortality; overexpression; pathogen; preference; pyruvate dehydrogenase; sugar; transcriptome

PROJECT SUMMARY The pathogenic protist Entamoeba histolytica is estimated to be responsible for 50,000 to 100,000 deaths each year and is a significant source of morbidity and mortality due to parasitic disease in humans, particularly in developing countries. This protist is also listed by NIAID as a category B priority pathogen with bioterrorism potential due to its low infectious dose and potential for dissemination through compromised food and water supplies. E. histolytica has a simple life cycle, existing in either an infectious cyst form or as motile trophozoites which can reside in the anaerobic confines of the human colon. This microbe is a scavenger that lacks ATP- producing mitochondria and relies on glycolysis as its primary pathway for generation of ATP. The PPi- dependent glycolytic pathway of Entamoeba has an increased net generation of ATP, which would be beneficial for organisms that lack a functional citric acid cycle and oxidative phosphorylation and must rely solely on substrate level phosphorylation to generate ATP from glucose breakdown. The proposed research examines the role of acetate fermentation in E. histolytica energy metabolism and physiology. Acetate is produced during growth in axenic culture, and the two enzymes of interest, acetate kinase (Ack) and ADP- forming acetyl-CoA synthetase (Acd), are both proposed to play a role in acetate production at the end of glycolysis. Acd is proposed to provide for generation of an additional ATP at the end of glycolysis. Recombinant E. histolytica Acd (EhAcd) has been shown to function well in both directions of the reaction, suggesting that this enzyme can operate in either direction in vivo, depending on the cell's metabolic needs. Recombinant E. histolytica Ack (EhAck) has been shown to have two unusual properties: production of PPi rather than ATP, and a strong preference for the acetate-forming direction of the reaction. Ack may thus provide a mechanism for supplying additional PPi for glycolysis under certain growth conditions that require a higher flux through this pathway. The two specific aims of this proposal are (1) to investigate the enzymology of EhAcd and EhAck using a structure-function approach; and (2) to determine the physiological roles of Acd and Ack in E. histolytica through transcriptome profiling and metabolic analysis of ACK and ACD RNAi and overexpression strains. The results from this research will provide a better understanding of a key metabolic pathway for ATP generation in this important eukaryotic pathogen.

项目总结