title: Force-dependent switch in protein unfolding pathways and transition-state movements.
abstract: Although it is known that single-domain proteins fold and unfold by parallel pathways, demonstration of this expectation has been difficult to establish in experiments. Unfolding rate, [Formula: see text], as a function of force f, obtained in single-molecule pulling experiments on src SH3 domain, exhibits upward curvature on a [Formula: see text] plot. Similar observations were reported for other proteins for the unfolding rate [Formula: see text]. These findings imply unfolding in these single-domain proteins involves a switch in the pathway as f or [Formula: see text] is increased from a low to a high value. We provide a unified theory demonstrating that if [Formula: see text] as a function of a perturbation (f or [Formula: see text]) exhibits upward curvature then the underlying energy landscape must be strongly multidimensional. Using molecular simulations we provide a structural basis for the switch in the pathways and dramatic shifts in the transition-state ensemble (TSE) in src SH3 domain as f is increased. We show that a single-point mutation shifts the upward curvature in [Formula: see text] to a lower force, thus establishing the malleability of the underlying folding landscape. Our theory, applicable to any perturbation that affects the free energy of the protein linearly, readily explains movement in the TSE in a β-sandwich (I27) protein and single-chain monellin as the denaturant concentration is varied. We predict that in the force range accessible in laser optical tweezer experiments there should be a switch in the unfolding pathways in I27 or its mutants.
authors.affiliation: Biophysics Program, Institute for Physical Science and Technology, Department of Chemistry & Biochemistry, University of Maryland, College Park, MD 20742; pavel.zhuravlev@gmail.com mxh605@case.edu.
authors.forename: Pavel I
authors.initials: PI
authors.lastname: Zhuravlev
authors.affiliation: Department of Physics, Case Western Reserve University, Cleveland, OH 44106; pavel.zhuravlev@gmail.com mxh605@case.edu.
authors.forename: Michael
authors.initials: M
authors.lastname: Hinczewski
authors.affiliation: Biophysics Program, Institute for Physical Science and Technology, Department of Chemistry & Biochemistry, University of Maryland, College Park, MD 20742;
authors.forename: Shaon
authors.initials: S
authors.lastname: Chakrabarti
authors.affiliation: California Institute for Quantitative Biosciences, Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720.
authors.forename: Susan
authors.initials: S
authors.lastname: Marqusee
authors.affiliation: Biophysics Program, Institute for Physical Science and Technology, Department of Chemistry & Biochemistry, University of Maryland, College Park, MD 20742;
authors.forename: D
authors.initials: D
authors.lastname: Thirumalai
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001696:Biomechanical Phenomena; D002645:Chickens; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009068:Movement; D050505:Mutant Proteins / Q000737:chemistry; D058767:Protein Unfolding; D013816:Thermodynamics; D018909:src Homology Domains
chemical_list: D050505:Mutant Proteins
keywords: parallel pathways; protein folding; single-molecule force spectroscopy
references.citation: Curr Opin Struct Biol. 2011 Feb;21(1):12-24
references.pmid: 21144739
references.citation: Proc Natl Acad Sci U S A. 1985 Jun;82(12):4028-30
references.pmid: 3858860
references.citation: J Mol Biol. 2005 Nov 11;353(5):1171-86
references.pmid: 16219323
references.citation: Curr Opin Struct Biol. 2007 Feb;17(1):21-9
references.pmid: 17251003
references.citation: Science. 2011 Oct 28;334(6055):512-6
references.pmid: 22034433
references.citation: J Mol Biol. 2014 Jul 15;426(14):2653-66
references.pmid: 24846645
references.citation: Biophys J. 2014 Jul 1;107(1):220-32
references.pmid: 24988356
references.citation: Science. 1997 May 16;276(5315):1109-12
references.pmid: 9148804
references.citation: J Chem Phys. 2011 Oct 28;135(16):164103
references.pmid: 22047224
references.citation: Proc Natl Acad Sci U S A. 2007 Dec 18;104(51):20268-73
references.pmid: 18079292
references.citation: Nature. 2003 May 8;423(6936):190-3
references.pmid: 12736689
references.citation: Nat Commun. 2015;6:6861
references.pmid: 25882479
references.citation: Proc Natl Acad Sci U S A. 1992 Sep 15;89(18):8721-5
references.pmid: 1528885
references.citation: Proc Natl Acad Sci U S A. 2014 Jun 24;111(25):9048-53
references.pmid: 24927549
references.citation: Science. 2014 Oct 31;346(6209):1254211
references.pmid: 25359979
references.citation: J Am Chem Soc. 2013 Oct 2;135(39):14768-74
references.pmid: 24015877
references.citation: Biophys J. 2006 May 15;90(10):3410-27
references.pmid: 16473903
references.citation: Proc Natl Acad Sci U S A. 1999 May 25;96(11):6166-70
references.pmid: 10339559
references.citation: Proteins. 1995 Mar;21(3):167-95
references.pmid: 7784423
references.citation: Proc Natl Acad Sci U S A. 2012 Oct 30;109(44):17820-5
references.pmid: 22949695
references.citation: Nature. 1999 Jan 7;397(6714):50-3
references.pmid: 9892352
references.citation: Nat Struct Biol. 2003 Aug;10(8):658-62
references.pmid: 12833152
references.citation: Proc Natl Acad Sci U S A. 2009 Nov 10;106(45):19011-6
references.pmid: 19887634
references.citation: J Chem Phys. 2012 Aug 7;137(5):055103
references.pmid: 22894385
references.citation: Proc Natl Acad Sci U S A. 2013 Mar 19;110(12):4500-5
references.pmid: 23487746
references.citation: Science. 2012 Apr 20;336(6079):362-6
references.pmid: 22517863
references.citation: Biopolymers. 1992 Jun;32(6):695-709
references.pmid: 1643270
references.citation: Curr Opin Struct Biol. 2004 Feb;14(1):76-88
references.pmid: 15102453
references.citation: Science. 1995 Mar 17;267(5204):1619-20
references.pmid: 7886447
references.citation: Proc Natl Acad Sci U S A. 2011 May 10;108(19):7787-92
references.pmid: 21512127
references.citation: Nature. 2003 May 8;423(6936):193-7
references.pmid: 12736690
references.citation: J Mol Biol. 2012 Jul 13;420(3):235-50
references.pmid: 22542529
references.citation: J Am Chem Soc. 2013 Aug 28;135(34):12722-9
references.pmid: 23905836