Multinuclear NMR studies of the flavodoxin from Anabaena 7120:. beta. -sheet structure and the flavin mononucleotide binding site
A concerted approach to primary {sup 1}H, {sup 13}C, and {sup 15}N nuclear magnetic resonance assignments in proteins was developed. The method requires enrichment of the protein with {sup 13}C and {sup 15}N. The technique relies on the comparison of data sets from NMR experiments that correlate various nuclei: {sup 13}C({sup 13}C) double quantum correlations, {sup 1}H({sup 13}C) and {sup 1}H({sup 15}N) single bond correlations, and {sup 1}H({sup 13}C) and {sup 1}H({sup 15}N) multiple bond correlations. Comparison of data sets increases the number of resonances that can be assigned and improves assignment confidence. By combined use of these and conventional NMR techniques, sequential assignments were made for the {beta}-sheet and flavin mononucleotide (FMN) binding site residues in flavodoxin from Anabaena 7120. The {beta}-sheet structure was found to be similar to that seen in the crystal structure of Anacystis nidulans flavodoxin. In the FMN binding site, a total of 69 NOEs were identified: eight between protons of FMN, 36 between protons of binding site residues, and 25 between protons of FMN and protein. These constraints were used to determine the localized solution structure of the flavin binding site. The electronic environment and conformation of the protein-bound isoalloxazine ring were investigated by determining chemical shifts and coupling constants for the ring atoms. The carbonyl edge of the flavin ring was found to be slightly polarized by hydrogen bonding to the protein. The xylene ring was found to be nonplanar. The C{sup 6}-N{sup 5} region of the flavin appears to be solvent accessible.
- Research Organization:
- Wisconsin Univ., Madison, WI (USA)
- OSTI ID:
- 6414695
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ACRIDINES
AMINES
AROMATICS
AZAARENES
AZINES
BARYONS
CARBON 13
CARBON ISOTOPES
ELEMENTARY PARTICLES
EVEN-ODD NUCLEI
FERMIONS
FLAVINES
HADRONS
HETEROCYCLIC COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
MAGNETIC RESONANCE
MEMBRANE PROTEINS
MOLECULAR STRUCTURE
NITROGEN 15
NITROGEN ISOTOPES
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
NUCLEI
NUCLEONS
NUCLEOTIDES
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PROTEINS
PROTONS
PYRIDINES
RECEPTORS
RESONANCE
SPECTRA
STABLE ISOTOPES
TRACER TECHNIQUES