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pH-dependent interactions and the stability and folding kinetics of the N-terminal domain of L9. Electrostatic interactions are only weakly formed in the transition state for folding.

Abstract：

:The role of electrostatic interactions in the stability and the folding of the N-terminal domain of the ribosomal protein L9 (NTL9) was investigated by determining the effects of varying the pH conditions. Urea denaturations and thermal unfolding experiments were used to measure the free energy of folding, DeltaG degrees, at 18 different pH values, ranging from pH 1.1 to pH 10.5. Folding rates were measured at 19 pH values between pH 2.1 and pH 9.5, and unfolding rates were determined at 15 pH values in this range using stopped-flow fluorescence experiments. The protein is maximally stable between pH 5.5 and 7.5 with a value of DeltaG degrees =4.45 kcal mol(-1). The folding rate reaches a maximum at pH 5.5, however the change in folding rates with pH is relatively modest. Over the pH range of 2.1 to 5.5 there is a small increase in folding rates, ln (k(f)) changes from 5.1 to 6.8. However, the change in stability is more dramatic, with a difference of 2.6 kcal mol(-1) between pH 2.0 and pH 5.4. The change in stability is largely due to the smaller barrier for unfolding at low pH values. The natural log of the unfolding rates varies by approximately four units between pH 2.1 and pH 5.5. The stability of the protein decreases above pH 7.5 and again the change is largely due to changes in the unfolding rate. ln (k(f)) varies by less than one unit between pH 5.5 and pH 9.5 while DeltaG degrees decreases by 2.4 kcal mol(-1) over the range of pH 5. 4 to pH 10.0, which corresponds to a change in ln K(eq) of 4.0. These studies show that pH-dependent interactions contribute significantly to the overall stability of the protein but have only a small effect upon the folding kinetics, indicating that electrostatic interactions are weakly formed in the transition state for folding.

journal_name

J Mol Biol

journal_title

Journal of molecular biology

authors

Luisi DL,Raleigh DP

doi

10.1006/jmbi.2000.3752

keywords:

subject

Has Abstract

pub_date

2000-06-16 00:00:00

pages

1091-100

issue

eissn

0022-2836

issn

1089-8638

pii

S0022-2836(00)93752-5

journal_volume

299

pub_type

杂志文章

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pH-dependent interactions and the stability and folding kinetics of the N-terminal domain of L9. Electrostatic interactions are only weakly formed in the transition state for folding.