Fluorescence studies with malate dehydrogenase from Bradyrhizobium japonicum 3I1B-143 bacteroids: a two-tryptophan containing protein.
A number of fluorescence studies, both of trp residues and bound NADH, have been reported for porcine malate dehydrogenase (MDH). The large number of trp residues (six) complicates the interpretation of some studies. To circumvent this we have performed studies with a two-tryptophan (per subunit) MDH from Bradyrhizobium japonicum 3I1B-143 bacteroids. We have performed phase/modulation fluorescence lifetime measurements, as a function of temperature and added quencher KI, in order to resolved the 1.2-ns (blue) and 6.5-ns (red) contributions from the two classes of trp residues. Anisotropy decay studies have also been performed. The binding of NADH dynamically quenches the fluorescence of both trp residues, but, unlike mammalian cytoplasmic and mitochondrial MDH, there is not a large enhancement in fluorescence of bound NADH upon forming a ternary complex with either tartronic acid or D-malate.
Ghiron
C A
CA
Department of Biochemistry, University of Missouri, Columbia 65211.
Eftink
M R
MR
Waters
J K
JK
Emerich
D W
DW
D007700:Kinetics; D007834:Lasers; D008291:Malate Dehydrogenase / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D011485:Protein Binding; D012230:Rhizobiaceae / Q000201:enzymology*; D013050:Spectrometry, Fluorescence / Q000379:methods; D014364:Tryptophan
D009243:NAD; D014364:Tryptophan; D008291:Malate Dehydrogenase