Titration of E. coli transhydrogenase domain III with bound NADP+ or NADPH studied by NMR reveals no pH-dependent conformational change in the physiological pH range.
Artikel i vetenskaplig tidskrift, 2005

A pH-titration 2D NMR study of Escherichia coli transhydrogenase domain III with bound NADP(+) or NADPH has been carried out, in which the pH was varied between 5.4 and 12. In this analysis, individual amide protons served as reporter groups. The apparent pK(a) values of the amide protons, determined from the pH-dependent chemical shift changes, were attributed to actual pK(a) values for several titrating residues in the protein. The essential Asp392 is shown to be protonated at neutral pH in both the NADP(+) and NADPH forms of domain III, but with a marked difference in pK(a) not only attributable to the charge difference between the substrates. Titrating residues found in loop D/alpha5 point to a conformational difference of these structural elements that is redox-dependent, but not pH dependent. The observed apparent pK(a) values of these residues are discussed in relation to the crystal structure of Rhodospirillum rubrum domain III, the solution structure of E. coli domain III and the mechanism of intact proton-translocating transhydrogenase.

chemistry

Protein Structure

Biomolecular

metabolism

Tertiary

Hydrogen-Ion Concentration

chemistry

Escherichia coli Proteins

NADP

Titrimetry

chemistry

chemistry

NADP Transhydrogenase

Nuclear Magnetic Resonance

Oxidation-Reduction

Protein Subunits

Författare

Anders Pedersen

Göteborgs universitet

Tomas Johansson

Chalmers, Kemi- och bioteknik

Jan Rydström

Göteborgs universitet

B Göran Karlsson

Chalmers, Kemi- och bioteknik

Biochimica et biophysica acta

0006-3002 (ISSN)

Vol. 1707 254-8

Ämneskategorier

Biokemi och molekylärbiologi

DOI

10.1016/j.bbabio.2004.12.004

PubMed

15863102