Locus of the catalytic sites of UDP-glucose dehydrogenase in the native enzyme hexamer as delineated by fluorescence energy transfer

Journal Article · Wed Oct 01 04:00:00 EDT 1980 · Biophys. J.; (United States)

DOI:https://doi.org/10.1016/S0006-3495(80)84940-X· OSTI ID:7032772

Franzen, J S ^[1]; Marchetti, P; Feingold, D S

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1. Univ. of Pittsburgh, PA

UDP-glucose dehydrogenase is comprised of six identical subunits arranged in a hexagonal manner. The half-sites reactivity behavior of the enzyme suggests that the array has 32 symmetry, i.e., it is a trimer of dimers. Since the catalytic site thiol groups are biphasically alkylated, one can prepare enzyme which has three catalytic sites covalently blocked with a fluorescent donor and three with an acceptor. Peptide mapping of tryptic digests of enzyme so modified shows that < 5% of the fluorophores are on noncatalytic site peptides. The AEDANS group has been used as a donor with either fluorescein, eosin, or nitrobenzoxadiazole as the acceptor. The extent of incorporation was determined from the radioactivity and/or optical absorbance of the modified protein. Energy transfer efficiencies were evaluated routinely from the size of donor fluorescence quenching, after it was certain that sensitized fluorescence data gave comparable results.

• OSTI ID: 7032772
• Journal Information: Biophys. J.; (United States), Journal Name: Biophys. J.; (United States) Vol. 32:1; ISSN BIOJA
• Country of Publication: United States
• Language: English

Related Subjects

550200* -- Biochemistry
551000 -- Physiological Systems
59 BASIC BIOLOGICAL SCIENCES
BIOPHYSICS
CARBOHYDRATES
CATALYSIS
CONFIGURATION
DIMERS
ENERGY TRANSFER
ENZYME ACTIVITY
ENZYMES
FLUORESCENCE
GLYCOSIDES
HEXAGONAL CONFIGURATION
LUMINESCENCE
MOLECULAR STRUCTURE
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
OXIDOREDUCTASES
STRUCTURAL CHEMICAL ANALYSIS
UDPG