| Autor: |
De Vitto H; The Hormel Institute, University of Minnesota, Austin, Minnesota 55912, United States., Belfon KKJ; Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, New York 11790, United States., Sharma N; The Hormel Institute, University of Minnesota, Austin, Minnesota 55912, United States., Toay S; Department of Biological Chemistry, Grinnell College, Grinnell, Iowa 50112, United States., Abendroth J; UCB BioSciences, Bainbridge Island, Washington 98110, United States.; Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, Washington 98104, United States., Dranow DM; UCB BioSciences, Bainbridge Island, Washington 98110, United States.; Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, Washington 98104, United States., Lukacs CM; UCB BioSciences, Bainbridge Island, Washington 98110, United States.; Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, Washington 98104, United States., Choi R; Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, Washington 98104, United States., Udell HS; Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, Washington 98104, United States., Willis S; Department of Chemistry, Rollins College, Winter Park, Florida 32789, United States., Barrera G; Department of Chemistry and Biochemistry, Weber State University, Ogden, Utah 84408, United States., Beyer O; Department of Chemistry and Biochemistry, University of Maryland, Baltimore County, Baltimore, Maryland 21250, United States., Li TD; Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, New York 11790, United States., Hicks KA; Chemistry Department, State University of New York at Cortland, Cortland, New York 13045, United States., Torelli AT; Department of Chemistry, Ithaca College, Ithaca, New York 14850, United States., French JB; The Hormel Institute, University of Minnesota, Austin, Minnesota 55912, United States. |
| Abstrakt: |
Thiamin and its phosphate derivatives are ubiquitous molecules involved as essential cofactors in many cellular processes. The de novo biosynthesis of thiamin employs the parallel synthesis of 4-methyl-5-(2-hydroxyethyl)thiazole (THZ-P) and 4-amino-2-methyl-5(diphosphooxymethyl) pyrimidine (HMP) pyrophosphate (HMP-PP), which are coupled to generate thiamin phosphate. Most organisms that can biosynthesize thiamin employ a kinase (HMPK or ThiD) to generate HMP-PP. In nearly all cases, this enzyme is bifunctional and can also salvage free HMP, producing HMP-P, the monophosphate precursor of HMP-PP. Here we present high-resolution crystal structures of an HMPK from Acinetobacter baumannii (AbHMPK), both unliganded and with pyridoxal 5-phosphate (PLP) noncovalently bound. Despite the similarity between HMPK and pyridoxal kinase enzymes, our kinetics analysis indicates that AbHMPK accepts HMP exclusively as a substrate and cannot turn over pyridoxal, pyridoxamine, or pyridoxine nor does it display phosphatase activity. PLP does, however, act as a weak inhibitor of AbHMPK with an IC 50 of 768 μM. Surprisingly, unlike other HMPKs, AbHMPK catalyzes only the phosphorylation of HMP and does not generate the diphosphate HMP-PP. This suggests that an additional kinase is present in A. baumannii , or an alternative mechanism is in operation to complete the biosynthesis of thiamin. |
