Protein 2D NMR analysis utilizing selective /sup 2/H and /sup 13/C enrichment

Conference · Thu May 01 00:00:00 EDT 1986 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:5282963

LeMaster, D M; Richards, F M

E. coli thioredoxin has been prepared with specific residue types substituted with fully enriched /sup 2/H or /sup 13/C labeled amino acids. In /sup 1/H COSY and NOESY experiments cross peaks result from pairs of protons interacting via through-bond or through-space coupling respectively. A cross peak is eliminated if either nucleus is substituted with deuterium. Direct residue type assignments of cross peaks have been obtained by comparing data from protein samples with one residue type deuterated and data from a non-enriched sample. Selective deuteration is particularly useful for the interresidue connectivity assignments obtained by NOESY experiments which normally depend on residue type assignment information obtained from independent COSY data. Difficulties in /sup 1/H COSY type intraresidue connectivity assignments are the main reason that successful analyses generally have been limited to proteins less than 10 kilodaltons. The authors have collected /sup 13/C homonuclear COSY data which compared to the /sup 1/H experiment benefit from larger spin couplings as well as from the spectral simplification obtained by enrichment. Resolved cross peaks for the /sup 13/C-/sup 13/C J/sub 1/ couplings are readily observed with 30 mg of labeled protein. /sup 1/H-/sup 13/C heteronuclear COSY experiments then provide an independent method of intraresidue proton spin connectivity assignments.

• Research Organization: Yale Univ., New Haven, CT
• OSTI ID: 5282963
• Report Number(s): CONF-8606151-
• Journal Information: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Vol. 45:6; ISSN FEPRA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
550600 -- Medicine
59 BASIC BIOLOGICAL SCIENCES
62 RADIOLOGY AND NUCLEAR MEDICINE
AMINO ACIDS
BACTERIA
BARYONS
CARBON 13
CARBON ISOTOPES
CARBOXYLIC ACIDS
DEUTERIUM
ELEMENTARY PARTICLES
ESCHERICHIA COLI
EVEN-ODD NUCLEI
FERMIONS
HADRONS
HYDROGEN ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
MAGNETIC RESONANCE
MICROORGANISMS
MOLECULAR STRUCTURE
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
NUCLEI
NUCLEONS
ODD-ODD NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
PROTEINS
PROTONS
RESONANCE
SPECTRA
STABLE ISOTOPES
TRACER TECHNIQUES