Proton NMR studies of lambda phage cro repressor
The cro repressor of bacteriophage lambda has been studied extensively in solution using two-dimensional nuclear magnetic resonance spectroscopy (2DNMR). Near-complete resonance assignments of the cro /sup 1/H NMR spectrum were obtained, using selectively optimized relayed coherence transfer spectroscopy (RELAY), and using the sequential resonance assignment method. The peptide backbone resonances for 65 of the 66 amino acids and most side chain resonances were assigned. The assignments helped identify over 300 NOEs occurring in the two-dimensional nuclear Overhauser effect spectroscopy (NOESY) spectrum, which were used to analyze the conformation of cro repressor in solution. The selectively optimized RELAY experiment was instrumental in identifying amino acid spin systems in the crowded cro /sup 1/H NMR spectrum. A mixing time of 26 msec yielded relatively strong C/sup ..cap alpha../H-C/sup ..cap alpha../H/sub 3/ RELAY cross peaks for Valine, Isoleucine, and Threonine residues, while RELAY cross peaks for other spin systems were weak or not observed. This promoted rapid and unambiguous identification of one-third of the spin systems in cro. The selectively optimized RELAY experiment should make it possible to investigate and assign and NMR spectra of proteins twice the size of cro repressor. With these spin systems identified, the ambiguities in spectral overlaps were reduced, and the sequential assignments could be completed. The NOEs identified in the cro spectrum were used in conjunction with distance geometry methods to calculate the conformation of ..cap alpha..2-..cap alpha..3 helices, which are involved in DNA recognition and binding.
- Research Organization:
- Washington Univ., Seattle (USA)
- OSTI ID:
- 6720581
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
62 RADIOLOGY AND NUCLEAR MEDICINE
AMINO ACIDS
BACTERIOPHAGES
BARYONS
CARBOXYLIC ACIDS
CONFORMATIONAL CHANGES
DNA
ELEMENTARY PARTICLES
FERMIONS
HADRONS
HYDROXY ACIDS
LEUCINE
MAGNETIC RESONANCE
MICROORGANISMS
MOLECULAR STRUCTURE
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
NUCLEIC ACIDS
NUCLEONS
ORGANIC ACIDS
ORGANIC COMPOUNDS
PARASITES
PEPTIDES
PROTEINS
PROTONS
RESONANCE
SPECTRA
THREONINE
VALINE
VIRUSES