PMID: 11910024Mar 23, 2002Paper

Analysis of serine proteinase-inhibitor interaction by alanine shaving

Protein Science : a Publication of the Protein Society
Olga BuczekJacek Otlewski

Abstract

We analyzed the energetic importance of residues surrounding the hot spot (the P(1) position) of bovine pancreatic trypsin inhibitor (BPTI) in interaction with two proteinases, trypsin and chymotrypsin, by a procedure called molecular shaving. One to eight residues of the structural epitope, composed of two extended and exposed loops, were mutated to alanine(s). Although truncation of the side chains of residues surrounding the P(1) position to methyl groups caused a decrease in Delta G(den) values up to 6.4 kcal mole(-1), it did not influence the overall conformation of the inhibitor. We found that the replacement of up to six residues with alanines was fully additive at the level of protein stability. To analyze the influence of the structural epitope on the association energy, we determined association constants for BPTI variants and both enzymes and applied the additivity analysis. Shaving of two binding loops led to a progressive drop in the association energy, more pronounced for trypsin (decrease up to 9.6 kcal mole(-1)) than chymotrypsin (decrease up to 3.5 kcal mole(-1)). In the case of extensively mutated variants interacting with chymotrypsin, the association energies agreed very well with the values calculated from ...Continue Reading

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Citations

Jan 17, 2004·Archives of Biochemistry and Biophysics·De-Yu XieRichard A Dixon
Feb 13, 2003·Current Opinion in Structural Biology·Michael LaskowskiZhengPing Yi
Jul 31, 2013·Quarterly Reviews of Biophysics·Alexey V Onufriev, Emil Alexov
Feb 23, 2008·Critical Reviews in Clinical Laboratory Sciences·Malgorzata ZakrzewskaAntoni Wiedlocha
Feb 8, 2006·Proteins·Zhengping YiMichael Laskowski
Dec 29, 2005·Journal of Molecular Biology·Dagmar Hobson, Olke C Uhlenbeck
Aug 30, 2005·Journal of Molecular Biology·Malgorzata ZakrzewskaJacek Otlewski
Jul 10, 2010·Biochimie·Peter GoettigHans Brandstetter
Apr 22, 2005·Journal of Biomolecular Structure & Dynamics·N Shamaladevi, Vasantha Pattabhi

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