This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Project I: Human pancreatic alpha-amylase (HPA;496 a.a.'s) is a critical digestive enzyme catalyzing the hydrolysis of starch and other long chain carbohydrates, which represent a major source of dietary glucose. A mechanistic understanding of this catalytic process would be of considerable importance in the treatment of diabetes and obesity, both chronic diseases exacting a heavy toll in term of health care outcomes. Future therapeutic development directed at HPA activity is largely dependent on a structural understanding of how catalytic residues of this enzyme function and the mode of substrate binding in the elongated binding cleft present. Although we have successfully applied site directed mutagenesis techniques and grown crystals of both wild-type and variant proteins, interpretation of the structural results of complexes formed by substrates and inhibitors has had limited success due to a lack of resolution (~1.9 ?) using our home laboratory x-ray source (Rigaku RU-300) and intend to extend the resolution to facilitate the development of novel therapeutics based on this mechanistic data. Project II: Surprisingly, only Gram-negative bacteria contain the hexameric Type II citrate synthases that have the special property of being metabolically regulated. In contrast, the dimeric citrate synthases of other organisms (including humans) is unregulated. Since many Gram-negative bacteria are dangerous human pathogens, the special properties of Type II citrate synthases could be a basis for the development of novel anti-microbials. Key to the development of such anti-microbials is the structural characterization of a Type II citrate synthase (CS;6 x 424 a.a.'s) in its various allosteric and metabolically controlled states. After considerable effort our laboratory has crystallized the E. coli enzyme and this remains the only such enzyme for which usable crystals have been obtained.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001209-32
Application #
8362109
Study Section
Special Emphasis Panel (ZRG1-BCMB-P (40))
Project Start
2011-03-01
Project End
2012-02-29
Budget Start
2011-03-01
Budget End
2012-02-29
Support Year
32
Fiscal Year
2011
Total Cost
$3,295
Indirect Cost
Name
Stanford University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
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