The effect of calciums on molecular motions of proteinase K.
The native serine protease proteinase K binds two calcium cations. It has been reported that Ca(2+) removal decreased the enzyme's thermal stability and to some extent the substrate affinity, but has discrepant effects on catalytic activity of the enzyme. Molecular dynamics simulations were performed on the Ca(2+)-bound and Ca(2+)-free proteases to investigate the mechanism by which the calciums affect the structural stability, molecular motions, and catalytic activity of proteinase K. Very similar structural properties were observed between these two forms of proteinase K during simulations; and several long-lived hydrogen bonds and salt bridges common to both forms of proteinase K were found to be crucial in maintaining the local conformations around these two Ca(2+) sites. Although Ca(2+) removal enhanced the overall flexibility of proteinase K, the flexibility in a limited number of segments surrounding the substrate-binding pockets decreased. The largest differences in the equilibrium structures of the two simulations indicate that, upon the removal of Ca(2+), the large concerted motion originating from the Ca1 site can transmit to the substrate-binding regions but not to the catalytic triad residues. In conjunction with the large overlap of the essential subspaces between the two simulations, these results not only provide insight into the dynamics of the underlying molecular mechanism responsible for the unchanged enzymatic activity as well as the decreased thermal stability and substrate affinity of proteinase K upon Ca(2+) removal, but also complement the experimentally determined structural and biochemical data.
Liu
Shu-Qun
SQ
Laboratory for Conservation and Utilization of Bio-Resources, Yunnan University, Kunming, Yunnan, People's Republic of China.
Tao
Yan
Y
Meng
Zhao-Hui
ZH
Fu
Yun-Xin
YX
Zhang
Ke-Qin
KQ
D001665:Binding Sites; D002118:Calcium / Q000737:chemistry*; D002384:Catalysis; D019286:Endopeptidase K / Q000737:chemistry*; D005656:Fungal Proteins / Q000737:chemistry; D006860:Hydrogen Bonding; D007700:Kinetics; D008958:Models, Molecular; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D011487:Protein Conformation; D013329:Structure-Activity Relationship
D005656:Fungal Proteins; D019286:Endopeptidase K; D002118:Calcium
Biochemistry. 2001 Mar 13;40(10):3080-8
11258922
Nature. 2007 Dec 13;450(7172):964-72
18075575
Nature. 1989 Feb 2;337(6206):481-4
2915693
Proc Natl Acad Sci U S A. 1958 Feb;44(2):98-104
16590179
FEBS J. 2008 Jan;275(1):117-27
18067583
Biophys J. 1999 Jul;77(1):505-15
10388775
Structure. 1997 Apr 15;5(4):521-32
9115441
J Biol Chem. 1991 Sep 15;266(26):17695-9
1894649
J Mol Biol. 1999 Feb 12;286(1):267-78
9931265
Biophys J. 2000 May;78(5):2191-200
10777720
Eur J Biochem. 1974 Aug 15;47(1):91-7
4373242
EMBO J. 1984 Jun;3(6):1311-4
6378621
Proteins. 1998 May 1;31(2):116-27
9593186
FEBS J. 2006 Jan;273(1):61-71
16367748
Protein Eng. 1997 Jan;10(1):31-7
9051731
Nature. 1989 May 18;339(6221):191-5
2716847
Anal Biochem. 1979 Sep 15;98(1):116-22
94516
J Mol Model. 2010 Jan;16(1):17-28
19466463
Proteins. 1993 Dec;17(4):412-25
8108382
J Mol Graph Model. 2007 Jul;26(1):306-18
17227719
FEBS Lett. 1986 Mar 3;197(1-2):105-10
3512298
J Biol Chem. 1994 Sep 16;269(37):23108-11
8083213
Eur J Biochem. 1988 Dec 1;178(1):155-71
3203685
J Mol Model. 2008 Sep;14(9):857-70
18594881
FEBS J. 2005 Feb;272(3):832-45
15670163
Protein Eng. 1991 Oct;4(7):719-37
1798697
Proc Natl Acad Sci U S A. 1998 May 26;95(11):5942-9
9600896
J Biol Chem. 2003 May 16;278(20):18434-9
12639952
Eur J Biochem. 1988 Sep 15;176(2):441-7
3166426
J Biol Chem. 2002 Sep 27;277(39):36678-88
12130645
BMC Struct Biol. 2007 May 18;7:33
17511867
Science. 2008 Jun 13;320(5882):1471-5
18556554
J Comput Chem. 2007 Sep;28(12):2075-84
17405124
Proteins. 1995 May;22(1):45-54
7675786
Protein Sci. 1997 Mar;6(3):501-23
9070434
Protein Eng. 1999 Sep;12(9):747-54
10506284
Biochem J. 1987 Aug 15;246(1):1-17
3314863