Irving, JA; Haq, I; Dickens, JA; Faull, SV; Lomas, DA; (2014) Altered native stability is the dominant basis for susceptibility of α1-antitrypsin mutants to polymerisation. Biochem J , 460 (1) pp. 103-115. 10.1042/BJ20131650.

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Abstract

Serpins are protease inhibitors whose most stable state is achieved upon transition of a central five-stranded β-sheet to a six-stranded form. Mutations, low pH, denaturants, and elevated temperatures promote this transition, which can result in a growing polymer chain of inactive molecules. Different types of polymer are possible, but experimentally, only heat has been shown to generate polymers in vitro consistent with ex vivo pathological specimens. Many mutations that alter the rate of heat-induced polymerisation have been described, but interpretation is problematic, because discrimination is lacking between the effect of global changes in native stability and specific effects on structural mechanism. We show that the temperature midpoint (Tm) of thermal denaturation reflects the transition of α1-antitrypsin to the polymerisation intermediate, and determine the relationship with fixed-temperature polymerisation half-times (t0.5) in the presence of stabilising additives (TMAO, sucrose and sodium sulphate), point mutations and disulphide bonds. Combined with a retrospective analysis of 31 mutants characterised in the literature, our data show that global changes to native state stability are the predominant basis for the effects of mutations and osmolytes on heat-induced polymerisation, summarised by the equation: ln(t0.5,mutant/t0.5,wild-type)=0.34×ΔTm. It is deviations from this relationship that hold key information about the polymerisation process.

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