Cofactor-Apoprotein Hydrogen Bonding in Oxidized and Fully Reduced Flavodoxin Monitored by Trans-Hydrogen-Bond Scalar Couplings

• Published in: Chembiochem : a European journal of chemical biology Vol. 5; no. 11; pp. 1523 - 1534
• Main Authors: Löhr, Frank, Yalloway, Gary N, Mayhew, Stephen G, Rüterjans, Heinz
• Format: Journal Article
• Language: English
• Published: Weinheim Wiley-VCH Verlag 05.11.2004; WILEY-VCH Verlag; WILEY‐VCH Verlag
• Subjects: Apoproteins - chemistry; Apoproteins - metabolism; Desulfovibrio vulgaris; Desulfovibrio vulgaris - metabolism; flavin; Flavodoxin - chemistry; Flavodoxin - metabolism; Hydrogen Bonding; Models, Molecular; Molecular Structure; NMR spectroscopy; Nuclear Magnetic Resonance, Biomolecular - methods; Oxidation-Reduction; Protein Binding; protein structures; quantitative J correlation; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Structure-Activity Relationship; TROSY
• ISSN: 1439-4227; 1439-7633; 1439-4227
• DOI: 10.1002/cbic.200400171

Hydrogen bonding plays a key role in the tight binding of the FMN cofactor and the regulation of its redox properties in flavodoxins. Hydrogen bonding interactions can be directly observed in solution by multidimensional heteronuclear NMR spectroscopy through the scalar couplings between donor and acceptor nuclei. Here we report on the detection of intermolecular trans-hydrogen-bond couplings (hJ) between the flavin ring system and the backbone of Desulfovibrio vulgaris flavodoxin in the oxidized and the two-electron reduced states. For this purpose, experiments are adapted from pulse sequences previously applied to determining hJ coupling constants in nucleic acid-base pairs and proteins. The resulting h²JN,N, h⁴JN,N, h³JC,N, and h¹JH,N couplings involve the ¹⁵N(1), ¹³C(2), and ¹⁵N(3) nuclei of the pyrimidine moiety of FMN, whereas no such interactions are detectable for ¹³C(4) and ¹⁵N(5). Several long-range ¹⁵N-¹⁵N, ¹³C-¹⁵N, and ¹H-¹⁵N J-coupling constants within the flavin are obtained as “by-products”. The magnitudes of both hJ and regular J couplings are found to be dependent on the redox state. In general, good correlations between hJ coupling constants and donor-group ¹H chemical shifts and also crystallographic donor-acceptor distances are observed.