sup 14,15 N, sup 13 C, sup 57 Fe, and sup 1,2 H Q-band ENDOR study of Fe-S proteins with clusters that have endogenous sulfur ligands

• Published in: Biochemistry (Easton) 1992-02, Vol.31:7
• Main Authors: Houseman, A.L.P., Chaoliang Fan, Werst, M.M., Hoffman, B.M., Byungha Oh, Markley, J.L., Kennedy, M.C., Beinert, H.
• Format: Article
• Language: English
• Subjects: 550601 - Medicine- Unsealed Radionuclides in Diagnostics; ALGAE; BARYONS; CARBON 13; CARBON ISOTOPES; DEUTERIUM COMPOUNDS; ELEMENTARY PARTICLES; ENDOR; EVEN-ODD NUCLEI; FERMIONS; FERREDOXIN; FREQUENCY RANGE; HADRONS; HYDROGEN COMPOUNDS; INTERMEDIATE MASS NUCLEI; IRON 57; IRON ISOTOPES; ISOTOPES; LIGHT NUCLEI; MAGNETIC RESONANCE; METALLOPROTEINS; MICROORGANISMS; MOLECULAR STRUCTURE; NITROGEN 14; NITROGEN 15; NITROGEN ISOTOPES; NUCLEI; NUCLEONS; ODD-EVEN NUCLEI; ODD-ODD NUCLEI; ORGANIC COMPOUNDS; PLANTS; PROTEINS; PROTONS; RADIOLOGY AND NUCLEAR MEDICINE; RESONANCE; STABLE ISOTOPES; UNICELLULAR ALGAE
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi00122a026

The benefits of performing ENDOR experiments at higher microwave frequency are demonstrated in a Q-band (35 GHz) ENDOR investigation of a number of proteins with (nFe-mS) clusters, n = 2,3,4. Each protein displays several resonances in the frequency range of 0-20 MHz. In all instances, features are seen near {nu} {approx} 13 and 8 MHz that can be assigned, respectively, to distant ENDOR from {sup 13}C in natural-abundance (1.1%) and from {sup 14}N. In addition, a number of proteins show local {sup 13}C ENDOR signals with resolved hyperfine interactions; these are assigned to the {beta} carbons of cysteines bound to the cluster. Quadrupole coupling constants are derived for both local and distant {sup 14}N signals. The interpretation of the data is supported by studies on {sup 15}N- and {sup 13}C-enriched ferredoxin (Fd) from Anabaena 7120, where the {sup 15}N signals can be clearly correlated with the corresponding {sup 14}N signals and where the {sup 13}C signals are strongly enhanced. Thus, the observation of {sup 14}N {Delta}m{sub I} = {plus minus} 2 signals at Q-band provides a new technique for examining weak interactions with a cluster. Six proteins show an additional pattern near {nu} {approx} 18 MHz that arises from {sup 57}Fe in natural abundance (2.2%) with A({sup 57}Fe) {approx} 36 MHz, which opens the possibility of studying proteins for which enrichment is impractical. Q-band ENDOR studies also have been carried out on four {sup 2}H-exchanged Fe-S proteins, and ENDOR detects exchangeable protons in each. The importance of these findings for the interpretation of X- and Q-band ENDOR at low radiofrequencies is discussed.