z-logo
Premium
Aggregation states of mitochondrial malate dehydrogenase
Author(s) -
Sánchez Susana A.,
Brunet Juan E.,
Hazlett Theodore L.,
Jameson David M.
Publication year - 1998
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560071016
Subject(s) - malate dehydrogenase , chemistry , mitochondrion , branched chain alpha keto acid dehydrogenase complex , biochemistry , physics , dehydrogenase , enzyme
The oligomeric state of fluorescein‐labeled mitochondrial malate dehydrogenase (L‐malate NAD + oxidoreductase; mMDH; EC 1.1.1.37), as a function of protein concentration, has been examined using steady‐state and dynamic polarization methodologies. A “global” rotational relaxation time of 103 ± 7 ns was found for micromolar concentrations of mMDH‐fluorescein, which is consistent with the reported size and shape of mMDH. Dilution of the mMDH‐fluorescein conjugates, prepared using a phosphate buffer protocol, to nanomolar concentrations had no significant effect on the rotational relaxation time of the adduct, indicating that the dimer‐monomer dissociation constant for mMDH is below 10 −9 M. In contrast to reports in the literature suggesting a pH‐dependent dissociation of mMDH, the oligomeric state of this mMDH‐fluorescein preparation remained unchanged between pH 5.0 and 8.0. Application of hydrostatic pressure up to 2.5 kilobars was ineffective in dissociating the mMDH dimer. However, the mMDH dimer was completely dissociated in 1.5 M guanidinium hydrochloride. Dilution of a mMDH‐fluorescein conjugate, prepared using a Tris buffer protocol, did show dissociation, which can be attributed to aggregates present in these preparations. These results are considered in light of the disparities in the literature concerning the properties of the mMDH dimer‐monomer equilibrium.

This content is not available in your region!

Continue researching here.

Having issues? You can contact us here