Relative stability of the open and closed conformations of the active site loop of streptavidin.
The eight-residue surface loop, 45-52 (Ser, Ala, Val, Gly, Asn, Ala, Glu, Ser), of the homotetrameric protein streptavidin has a "closed" conformation in the streptavidin-biotin complex, where the corresponding binding affinity is one of the strongest found in nature (ΔG ∼ -18 kcal∕mol). However, in most of the crystal structures of apo (unbound) streptavidin, the loop conformation is "open" and typically exhibits partial disorder and high B-factors. Thus, it is plausible to assume that the loop structure is changed from open to closed upon binding of biotin, and the corresponding difference in free energy, ΔF = F(open) - F(closed) in the unbound protein, should therefore be considered in the total absolute free energy of binding. ΔF (which has generally been neglected) is calculated here using our "hypothetical scanning molecular-dynamics" (HSMD) method. We use a protein model in which only the atoms closest to the loop are considered (the "template") and they are fixed in the x-ray coordinates of the free protein; the x-ray conformation of the closed loop is attached to the same (unbound) template and both systems are capped with the same sphere of TIP3P water. Using the force field of the assisted model building with energy refinement (AMBER), we carry out two separate MD simulations (at temperature T = 300 K), starting from the open and closed conformations, where only the atoms of the loop and water are allowed to move (the template-water and template-loop interactions are considered). The absolute F(open) and F(closed) (of loop + water) are calculated from these trajectories, where the loop and water contributions are obtained by HSMD and a thermodynamic integration (TI) process, respectively. The combined HSMD-TI procedure leads to total (loop + water) ΔF = -27.1 ± 2.0 kcal∕mol, where the entropy TΔS constitutes 34% of ΔF, meaning that the effect of S is significant and should not be ignored. Also, ΔS is positive, in accord with the high flexibility of the open loop observed in crystal structures, while the energy ΔE is unexpectedly negative, thus also adding to the stability of the open loop. The loop and the 250 capped water molecules are the largest system studied thus far, which constitutes a test for the efficiency of HSMD-TI; this efficiency and technical issues related to the implementation of the method are also discussed. Finally, the result for ΔF is a prediction that will be considered in the calculation of the absolute free energy of binding of biotin to streptavidin, which constitutes our next project.
Department of Computational and Systems Biology, University of Pittsburgh School of Medicine, 3059 BST3, Pittsburgh, Pennsylvania 15260, USA.
Ignacio J
IJ
General
Hagai
H
Meirovitch
D001710:Biotin / Q000737:chemistry; D020134:Catalytic Domain; D056004:Molecular Dynamics Simulation; D011487:Protein Conformation; D019809:Streptavidin / Q000737:chemistry*; D013816:Thermodynamics
D001710:Biotin; D019809:Streptavidin
Proc Natl Acad Sci U S A. 1989 Apr;86(7):2190-4
2928324
Methods Enzymol. 1990;184:49-51
2201883
Curr Med Chem. 2006;13(29):3583-608
17168725
J Phys Chem B. 2009 Jun 4;113(22):7950-64
19435302
J Chem Phys. 2005 Dec 1;123(21):214908
16356071
J Chem Theory Comput. 2008 Jan;4(1):216
26619994
Arch Biochem Biophys. 1964 Jul 20;106:1-5
14217155
Science. 1992 Feb 21;255(5047):959-65
1546293
Nucleic Acids Res. 2006 Jul 1;34(Web Server issue):W24-31
16845002
J Phys Chem B. 2005 Nov 24;109(46):21963-70
16853854
Entropy (Basel). 2010 Aug 25;12(8):1946-1974
21448250
Proteins. 2010 May 15;78(7):1705-23
20186976
J Phys Chem B. 2008 Aug 7;112(31):9512-22
18613721
Proteins. 2002 May 1;47(2):194-208
11933066
Science. 1987 Mar 6;235(4793):1191-6
3823879
J Chem Phys. 2006 May 28;124(20):204108
16774320
Science. 1987 Jan 16;235(4786):318-21
3798113
Nature. 1977 Jun 16;267(5612):585-90
301613
Science. 1984 Sep 7;225(4666):983-9
17783020
J Phys Chem B. 2009 May 14;113(19):6971-85
19374419
Biochemistry. 1992 Oct 6;31(39):9350-4
1390720
J Phys Chem B. 2012 Jun 14;116(23):6628-36
22300239
J Chem Theory Comput. 2008 Jan;4(1):192-208
26619992
J Chem Phys. 2005 Feb 1;122(5):54903
15740349
Chem Rev. 2009 Sep;109(9):4092-107
19588959
Annu Rev Biophys Biophys Chem. 1989;18:431-92
2660832
Curr Opin Struct Biol. 2007 Apr;17(2):181-6
17395451
Phys Rev A Gen Phys. 1985 Dec;32(6):3709-3715
9896540
J Chem Phys. 2006 Jul 14;125(2):24905
16848609
Macromolecules. 1976 May-Jun;9(3):395-407
940353
J Chem Phys. 2004 Dec 8;121(22):10889-904
15634040
Angew Chem Int Ed Engl. 2006 Jun 19;45(25):4064-92
16761306
Protein Sci. 1998 Apr;7(4):848-59
9568892
Biochemistry. 1998 Jun 2;37(22):7965-80
9609689
Adv Protein Chem. 1975;29:85-133
237414
J Chem Theory Comput. 2005 Sep;1(5):1017-28
26641917
Science. 1989 Jan 6;243(4887):85-8
2911722
Biophys J. 1997 Mar;72(3):1047-69
9138555
Protein Sci. 1997 Jun;6(6):1157-66
9194176
Proteins. 2004 Dec 1;57(4):645-50
15481087
Proteins. 2010 May 15;78(7):1724-35
20186973