): Mechanisms of enzyme catalysis and regulation at the molecular level are issues fundamental to contemporary biology. Proposed are investigations of two highly regulated enzymes of carbohydrate metabolism, human brain hexokinase and porcine liver fructose-1,6-bisphosphatase. Brain hexokinase (Glucose + MgATP2- = Glucose-6-P2- + MgADP1- +H+) is the """"""""pacemaker of glycolysis"""""""" in brain tissue and the red blood cell. Fructose 1,6-bisphosphatue (fructose 1,6-bisphosphate + H2O = fructose-6-P + phosphate) is a key, regulatory enzyme in gluconeogenesis. Pig kidney fructose 1,6-bisphosphatase has been the focus of numerous crystallographic studies. Both of these enzymes have been focal points of the research during the past 30 years; however, because of the paucity of brain hexokinase in brain tissue, most investigations until recently have centered on fructose-1,6-bisphosphatase which is present in liver tissue in relatively large quantities. The group recently cloned, expressed (in Escherichia coli), and crystallized human brain hexokinase and porcine liver fructose-1,6-bisphosphatase (identical in amino acid sequence to the kidney enzyme). Thus, essential prerequisites for modern-day structure/function investigations such as the availability of cloned genes, expression systems, large quantities of pure enzyme and crystals, are now in place. The focus here is studies in directed mutation, kinetics, NMR and crystallography as a means of defining the atomic-level mechanisms of catalysis and regulation of fructose-1,6-bisphosphatase and brain hexokinase.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
3R01NS010546-34S1
Application #
6323805
Study Section
Physiological Chemistry Study Section (PC)
Program Officer
Jacobs, Tom P
Project Start
1975-09-01
Project End
2002-04-30
Budget Start
2000-05-01
Budget End
2001-04-30
Support Year
34
Fiscal Year
2000
Total Cost
$50,000
Indirect Cost
Name
Iowa State University
Department
Biochemistry
Type
Other Domestic Higher Education
DUNS #
City
Ames
State
IA
Country
United States
Zip Code
50011
Gao, Yang; Shen, Lu; Honzatko, Richard B (2014) Central cavity of fructose-1,6-bisphosphatase and the evolution of AMP/fructose 2,6-bisphosphate synergism in eukaryotic organisms. J Biol Chem 289:8450-61
Gao, Yang; Iancu, Cristina V; Mukind, Susmith et al. (2013) Mechanism of displacement of a catalytically essential loop from the active site of mammalian fructose-1,6-bisphosphatase. Biochemistry 52:5206-16
Zhou, Ke; Gao, Yang; Hoy, Julie A et al. (2012) Insights into diterpene cyclization from structure of bifunctional abietadiene synthase from Abies grandis. J Biol Chem 287:6840-50
Joseph, Raji E; Ginder, Nathaniel D; Hoy, Julie A et al. (2012) Structure of the interleukin-2 tyrosine kinase Src homology 2 domain; comparison between X-ray and NMR-derived structures. Acta Crystallogr Sect F Struct Biol Cryst Commun 68:145-53
Gao, Yang; Honzatko, Richard B; Peters, Reuben J (2012) Terpenoid synthase structures: a so far incomplete view of complex catalysis. Nat Prod Rep 29:1153-75
Joseph, Raji E; Ginder, Nathaniel D; Hoy, Julie A et al. (2011) Purification, crystallization and preliminary crystallographic analysis of the SH2 domain of IL-2-inducible T-cell kinase. Acta Crystallogr Sect F Struct Biol Cryst Commun 67:269-73
Leung, Daisy W; Borek, Dominika; Farahbakhsh, Mina et al. (2010) Crystallization and preliminary X-ray analysis of Ebola VP35 interferon inhibitory domain mutant proteins. Acta Crystallogr Sect F Struct Biol Cryst Commun 66:689-92
Leung, Daisy W; Prins, Kathleen C; Borek, Dominika M et al. (2010) Structural basis for dsRNA recognition and interferon antagonism by Ebola VP35. Nat Struct Mol Biol 17:165-72
Warner, Christopher D; Hoy, Julie A; Shilling, Taran C et al. (2010) Tertiary structure and characterization of a glycoside hydrolase family 44 endoglucanase from Clostridium acetobutylicum. Appl Environ Microbiol 76:338-46
Leung, Daisy W; Ginder, Nathaniel D; Fulton, D Bruce et al. (2009) Structure of the Ebola VP35 interferon inhibitory domain. Proc Natl Acad Sci U S A 106:411-6

Showing the most recent 10 out of 11 publications