pH-Induced denaturation of proteins: A single salt bridge contributes 3-5 kcal/mol to the free energy of folding of T4 lysozyme
- Univ. of Oregon, Eugene (USA)
The energetics of a salt bridge formed between the side chains of aspartic acid 70 (Asp70) and histidine 31 (His31) of T4 lysozyme have been examined by nuclear magnetic resonance techniques. The pK{sub a} values of the residues in the native state are perturbed from their values in the unfolded protein such that His31 has a pK{sub a} value of 9.1 in the native state and 6.8 in the unfolded state at 10{degree}C in moderate salt. Similarly, the aspartate pK{sub a} is shifted to a value of about 0.5 in the native state from its value of 3.5-4.0 in the unfolded state. These shifts in pK{sub a} show that the salt bridge is stabilized 3-5 kcal/mol. These observations and consideration of the thermodynamic coupling of protonation state to folding of proteins suggest a mechanism of acid denaturation in which the unfolded state is progressively stabilized by protonation of its acid residues as pH is lowered below pH 4. The unfolded state is stabilized only if acidic groups in the folded state have lower pK{sub a} values than in the unfolded state. When the pH is sufficiently low, the acid groups of both the native and unfolded states are fully protonated, and the apparent unfolding equilibrium constant becomes pH independent. Similar arguments apply to base-induced unfolding. These observations suggest that the electrostatic contribution of each ionizable group to the stability of the folded state can be directly assessed by simply measuring its apparent pK{sub a} by NMR or other methods.
- OSTI ID:
- 7192960
- Journal Information:
- Biochemistry; (USA), Vol. 29:9; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ASPARTIC ACID
NUCLEAR MAGNETIC RESONANCE
HISTIDINE
LYSOZYME
MOLECULAR MODELS
BACTERIOPHAGES
BIOCHEMICAL REACTION KINETICS
CARBON 13
CHEMICAL SHIFT
MUTANTS
PH VALUE
PROTEIN DENATURATION
X-RAY DIFFRACTION
AMINO ACIDS
AZOLES
CARBON ISOTOPES
CARBOXYLIC ACIDS
COHERENT SCATTERING
DIFFRACTION
ENZYMES
EVEN-ODD NUCLEI
GLYCOSYL HYDROLASES
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
HYDROLASES
IMIDAZOLES
ISOTOPES
KINETICS
LIGHT NUCLEI
MAGNETIC RESONANCE
MATHEMATICAL MODELS
MICROORGANISMS
NUCLEI
O-GLYCOSYL HYDROLASES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PARASITES
REACTION KINETICS
RESONANCE
SCATTERING
STABLE ISOTOPES
VIRUSES
550601* - Medicine- Unsealed Radionuclides in Diagnostics