Barstar mutant

The contribution of electrostatic interactions to fast association was analyzed by applying the classical Debye-Hiickel theory of electrostatic interactions between ions to mutants of bamase and barstar whose ionic side chains had been altered by protein engineering (Chapter 14).16 The association fits a two-step model that is probably general (equation 4.84). 

Interactions Analysis of the Barnase-Barstar Interface by Single Mutations and Double Mutant Cycles. 

Schreiber G, Fersht AR (1995) Energetics of protein-protein interactions analysis of the barnase-barstar interface by single mutations and double mutant cycles, J Mol Biol, 248 -178 1X9... 

Wild type barstar shows two fluorescence lifetimes, 4.1 and 1.5 ns. The long lifetime is attributed to two of the three Trp residues including Trp53. Thus, trcaiiparing lifetime d of void type and of double mutants, one found difficult any inteipreiation of the results with the conforniers model, especially if we condfkr tiiat the tertiary structure of barstar is identical in the mutant and wild type fbnns. 

The association of bamase, an extracellular ribonuclease, with its intracellular inhibitor, barstar, provides a particularly well-characterized example of electrostatically steered protein-protein encounter. The association rate is very fast (about 10 -10 at 50 mM ionic strength), and mutation and ionic-strength-dependence studies clearly show the influence of electrostatic interactions. Brownian dynamics simulations are able to reproduce the ionic-strength dependence of the rate for the wild-type proteins and the rates for wild-type and 11 mutants at 50 mM ionic strength to within a factor of 2. These simulations provide insight into the structure of the encounter (transition state) complex in which barstar tends to be shifted from its position in the bound complex towards the guanine binding loop on bamase. 

Fig. 3.2. From top to bottom Complexes of barstar with the mutant peptide inhibitor barstar-C40A/C82A) (2.2) and with d(CGAC) (2.3).

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