Carbonyl /sup 13/C NMR spectrum of basin pancreatic trypsin inhibitor: resonance assignments by selective amide hydrogen isotope labeling and detection of isotope effects on /sup 13/C nuclear shielding
The carbonyl region of the natural abundance /sup 13/C nuclear magnetic resonance (NMR) spectrum of basic pancreatic trypsin inhibitor is examined, and 65 of the 66 expected signals are characterized at varying pH and temperature. Assignments are reported for over two-thirds of the signals, including those of all buried backbone amide groups with slow proton exchange and all side-chain carbonyl groups. This is the first extensively assigned carbonyl spectrum for any protein. A method for carbonyl resonance assignments utilizing amide proton exchange and isotope effects on nuclear shielding is described in detail. The assignments are made by establishing kinetic correlation between effects of amide proton exchange observed in the carbonyl /sup 13/C region with development of isotope effects and in the amide proton region with disappearance of preassigned resonances. Several aspects of protein structure and dynamics in solution may be investigated by carbonyl /sup 13/C NMR spectroscopy. Some effects of side-chain primary amide group hydrolysis are described. The main interest is on information about intramolecular hydrogen-bond energies and changes in the protein due to amino acid replacements by chemical modification or genetic engineering.
- Research Organization:
- Univ. of Roskilde (Denmark)
- OSTI ID:
- 6243529
- Journal Information:
- Biochemistry; (United States), Vol. 27:23
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ENZYME INHIBITORS
NUCLEAR MAGNETIC RESONANCE
TRYPSIN
BIOCHEMICAL REACTION KINETICS
CARBON 13
DEUTERIUM COMPOUNDS
ISOTOPE EFFECTS
NMR SPECTRA
OVERHAUSER EFFECT
PROTEIN STRUCTURE
CARBON ISOTOPES
ENZYMES
EVEN-ODD NUCLEI
HYDROGEN COMPOUNDS
HYDROLASES
ISOTOPES
KINETICS
LIGHT NUCLEI
MAGNETIC RESONANCE
NUCLEI
PEPTIDE HYDROLASES
REACTION KINETICS
RESONANCE
SERINE PROTEINASES
SPECTRA
STABLE ISOTOPES
550601* - Medicine- Unsealed Radionuclides in Diagnostics