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PDBsum entry 2snm
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* Residue conservation analysis
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Enzyme class:
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E.C.3.1.31.1
- micrococcal nuclease.
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Reaction:
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Endonucleolytic cleavage to nucleoside 3'-phosphates and 3'-phosphooligonucleotide end-products.
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J Mol Biol
221:7
(1991)
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PubMed id:
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In a staphylococcal nuclease mutant the side-chain of a lysine replacing valine 66 is fully buried in the hydrophobic core.
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W.E.Stites,
A.G.Gittis,
E.E.Lattman,
D.Shortle.
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ABSTRACT
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The crystal structure of the staphylococcal nuclease mutant V66K, in which
valine 66 is replaced by lysine, has been solved at 1.97 A resolution. Unlike
lysine residues in previously reported protein structures, this residue appears
to bury its side-chain in the hydrophobic core without salt bridging, hydrogen
bonding or other forms of electrostatic stabilization. Solution studies of the
free energy of denaturation, delta GH2O, show marked pH dependence and clearly
indicate that the lysine residue must be deprotonated in the folded state. V66K
is highly unstable at neutral pH but only modestly less stable than the
wild-type protein at high pH. The pH dependence of stability for V66K, in
combination with similar measurements for the wild-type protein, allowed
determination of the pKa values of the lysine in both the denatured and native
forms. The epsilon-amine of this residue has a pKa value in the denatured state
of 10.2, but in the native state it must be 6.4 or lower. The epsilon-amine is
thus deprotonated in the folded molecule. These values enabled an estimation of
the epsilon-amine's relative change in free energy of solvation between solvent
and the protein interior at 5.1 kcal/mol or greater. This implies that the value
of the dielectric constant of the protein interior must be less than 12.8.
Lysine is usually found with the methylene groups of its side-chain partly
buried but is nevertheless considered a hydrophilic surface residue. It would
appear that the high pKa value of lysine, which gives it a positive charge at
physiological pH, is the primary reason for its almost exclusive confinement to
the surface proteins. When deprotonated, this amino acid type can be fully
incorporated into the hydrophobic core.
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Literature references that cite this PDB file's key reference
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PubMed id
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Reference
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D.G.Isom,
C.A.Castañeda,
B.R.Cannon,
and
B.García-Moreno
(2011).
Large shifts in pKa values of lysine residues buried inside a protein.
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Proc Natl Acad Sci U S A,
108,
5260-5265.
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D.G.Isom,
C.A.Castañeda,
B.R.Cannon,
P.D.Velu,
and
B.García-Moreno E
(2010).
Charges in the hydrophobic interior of proteins.
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Proc Natl Acad Sci U S A,
107,
16096-16100.
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C.Carrasco-López,
C.Godoy,
B.de Las Rivas,
G.Fernández-Lorente,
J.M.Palomo,
J.M.Guisán,
R.Fernández-Lafuente,
M.Martínez-Ripoll,
and
J.A.Hermoso
(2009).
Activation of bacterial thermoalkalophilic lipases is spurred by dramatic structural rearrangements.
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J Biol Chem,
284,
4365-4372.
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PDB code:
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F.Milletti,
L.Storchi,
and
G.Cruciani
(2009).
Predicting protein pK(a) by environment similarity.
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Proteins,
76,
484-495.
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G.Xu,
S.B.Shin,
and
S.R.Jaffrey
(2009).
Global profiling of protease cleavage sites by chemoselective labeling of protein N-termini.
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Proc Natl Acad Sci U S A,
106,
19310-19315.
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M.C.Ho,
J.F.Ménétret,
H.Tsuruta,
and
K.N.Allen
(2009).
The origin of the electrostatic perturbation in acetoacetate decarboxylase.
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Nature,
459,
393-397.
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PDB codes:
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M.Sagermann,
R.R.Chapleau,
E.DeLorimier,
and
M.Lei
(2009).
Using affinity chromatography to engineer and characterize pH-dependent protein switches.
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Protein Sci,
18,
217-228.
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PDB codes:
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D.G.Isom,
B.R.Cannon,
C.A.Castañeda,
A.Robinson,
and
B.García-Moreno
(2008).
High tolerance for ionizable residues in the hydrophobic interior of proteins.
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Proc Natl Acad Sci U S A,
105,
17784-17788.
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J.L.Schlessman,
C.Abe,
A.Gittis,
D.A.Karp,
M.A.Dolan,
and
B.García-Moreno E
(2008).
Crystallographic study of hydration of an internal cavity in engineered proteins with buried polar or ionizable groups.
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Biophys J,
94,
3208-3216.
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PDB codes:
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L.Mitra,
J.B.Rouget,
B.Garcia-Moreno,
C.A.Royer,
and
R.Winter
(2008).
Towards a quantitative understanding of protein hydration and volumetric properties.
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Chemphyschem,
9,
2715-2721.
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M.J.Harms,
J.L.Schlessman,
M.S.Chimenti,
G.R.Sue,
A.Damjanović,
and
B.García-Moreno
(2008).
A buried lysine that titrates with a normal pKa: role of conformational flexibility at the protein-water interface as a determinant of pKa values.
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Protein Sci,
17,
833-845.
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PDB code:
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M.Schmidt Am Busch,
A.Lopes,
D.Mignon,
and
T.Simonson
(2008).
Computational protein design: software implementation, parameter optimization, and performance of a simple model.
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J Comput Chem,
29,
1092-1102.
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Q.Wang,
F.Cheng,
M.Lu,
X.Tian,
and
J.Ma
(2008).
Crystal structure of unliganded influenza B virus hemagglutinin.
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J Virol,
82,
3011-3020.
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PDB code:
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D.A.Karp,
A.G.Gittis,
M.R.Stahley,
C.A.Fitch,
W.E.Stites,
and
B.García-Moreno E
(2007).
High apparent dielectric constant inside a protein reflects structural reorganization coupled to the ionization of an internal Asp.
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Biophys J,
92,
2041-2053.
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PDB code:
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D.M.LeMaster,
J.S.Anderson,
and
G.Hernández
(2007).
Spatial distribution of dielectric shielding in the interior of Pyrococcus furiosus rubredoxin as sampled in the subnanosecond timeframe by hydrogen exchange.
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Biophys Chem,
129,
43-48.
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R.J.Johnson,
S.R.Lin,
and
R.T.Raines
(2007).
Genetic selection reveals the role of a buried, conserved polar residue.
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Protein Sci,
16,
1609-1616.
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A.Damjanović,
B.García-Moreno,
E.E.Lattman,
and
A.E.García
(2005).
Molecular dynamics study of water penetration in staphylococcal nuclease.
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Proteins,
60,
433-449.
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B.Kuhn,
P.A.Kollman,
and
M.Stahl
(2004).
Prediction of pKa shifts in proteins using a combination of molecular mechanical and continuum solvent calculations.
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J Comput Chem,
25,
1865-1872.
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H.Feng,
and
Y.Bai
(2004).
Repacking of hydrophobic residues in a stable mutant of apocytochrome b562 selected by phage-display and proteolysis.
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Proteins,
56,
426-429.
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N.Pokala,
and
T.M.Handel
(2004).
Energy functions for protein design I: efficient and accurate continuum electrostatics and solvation.
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Protein Sci,
13,
925-936.
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V.P.Denisov,
J.L.Schlessman,
B.García-Moreno E,
and
B.Halle
(2004).
Stabilization of internal charges in a protein: water penetration or conformational change?
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Biophys J,
87,
3982-3994.
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PDB code:
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K.Lim,
A.Tempczyk,
N.Bonander,
J.Toedt,
A.Howard,
E.Eisenstein,
and
O.Herzberg
(2003).
A catalytic mechanism for D-Tyr-tRNATyr deacylase based on the crystal structure of Hemophilus influenzae HI0670.
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J Biol Chem,
278,
13496-13502.
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PDB code:
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P.Pattanaik,
G.Ravindra,
C.Sengupta,
K.Maithal,
P.Balaram,
and
H.Balaram
(2003).
Unusual fluorescence of W168 in Plasmodium falciparum triosephosphate isomerase, probed by single-tryptophan mutants.
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Eur J Biochem,
270,
745-756.
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S.Balaji,
S.Aruna,
and
N.Srinivasan
(2003).
Tolerance to the substitution of buried apolar residues by charged residues in the homologous protein structures.
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Proteins,
53,
783-791.
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C.A.Fitch,
D.A.Karp,
K.K.Lee,
W.E.Stites,
E.E.Lattman,
and
B.García-Moreno E
(2002).
Experimental pK(a) values of buried residues: analysis with continuum methods and role of water penetration.
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Biophys J,
82,
3289-3304.
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K.K.Lee,
C.A.Fitch,
J.T.Lecomte,
and
B.García-Moreno E
(2002).
Electrostatic effects in highly charged proteins: salt sensitivity of pKa values of histidines in staphylococcal nuclease.
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Biochemistry,
41,
5656-5667.
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A.Ababou,
and
J.R.Desjarlais
(2001).
Solvation energetics and conformational change in EF-hand proteins.
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Protein Sci,
10,
301-312.
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C.N.Schutz,
and
A.Warshel
(2001).
What are the dielectric "constants" of proteins and how to validate electrostatic models?
|
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Proteins,
44,
400-417.
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V.V.Loladze,
D.N.Ermolenko,
and
G.I.Makhatadze
(2001).
Heat capacity changes upon burial of polar and nonpolar groups in proteins.
|
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Protein Sci,
10,
1343-1352.
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A.M.Lambeir,
J.Backmann,
J.Ruiz-Sanz,
V.Filimonov,
J.E.Nielsen,
I.Kursula,
B.V.Norledge,
and
R.K.Wierenga
(2000).
The ionization of a buried glutamic acid is thermodynamically linked to the stability of Leishmania mexicana triose phosphate isomerase.
|
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Eur J Biochem,
267,
2516-2524.
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PDB code:
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C.M.Dupureur,
and
L.H.Conlan
(2000).
A catalytically deficient active site variant of PvuII endonuclease binds Mg(II) ions.
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Biochemistry,
39,
10921-10927.
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J.J.Dwyer,
A.G.Gittis,
D.A.Karp,
E.E.Lattman,
D.S.Spencer,
W.E.Stites,
and
B.García-Moreno E
(2000).
High apparent dielectric constants in the interior of a protein reflect water penetration.
|
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Biophys J,
79,
1610-1620.
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K.K.Reiling,
T.R.Pray,
C.S.Craik,
and
R.M.Stroud
(2000).
Functional consequences of the Kaposi's sarcoma-associated herpesvirus protease structure: regulation of activity and dimerization by conserved structural elements.
|
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Biochemistry,
39,
12796-12803.
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PDB code:
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M.R.Gunner,
M.A.Saleh,
E.Cross,
A.ud-Doula,
and
M.Wise
(2000).
Backbone dipoles generate positive potentials in all proteins: origins and implications of the effect.
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Biophys J,
78,
1126-1144.
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T.Kajander,
P.C.Kahn,
S.H.Passila,
D.C.Cohen,
L.Lehtiö,
W.Adolfsen,
J.Warwicker,
U.Schell,
and
A.Goldman
(2000).
Buried charged surface in proteins.
|
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Structure,
8,
1203-1214.
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PDB code:
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B.Gopal,
S.S.Ray,
R.S.Gokhale,
H.Balaram,
M.R.Murthy,
and
P.Balaram
(1999).
Cavity-creating mutation at the dimer interface of Plasmodium falciparum triosephosphate isomerase: restoration of stability by disulfide cross-linking of subunits.
|
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Biochemistry,
38,
478-486.
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E.G.Alexov,
and
M.R.Gunner
(1999).
Calculated protein and proton motions coupled to electron transfer: electron transfer from QA- to QB in bacterial photosynthetic reaction centers.
|
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Biochemistry,
38,
8253-8270.
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M.H.Cordes,
N.P.Walsh,
C.J.McKnight,
and
R.T.Sauer
(1999).
Evolution of a protein fold in vitro.
|
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Science,
284,
325-328.
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PDB code:
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C.Lee,
M.N.Liang,
K.M.Tate,
J.D.Rabinowitz,
C.Beeson,
P.P.Jones,
and
H.M.McConnell
(1998).
Evidence that the autoimmune antigen myelin basic protein (MBP) Ac1-9 binds towards one end of the major histocompatibility complex (MHC) cleft.
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J Exp Med,
187,
1505-1516.
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E.J.Hebert,
A.Giletto,
J.Sevcik,
L.Urbanikova,
K.S.Wilson,
Z.Dauter,
and
C.N.Pace
(1998).
Contribution of a conserved asparagine to the conformational stability of ribonucleases Sa, Ba, and T1.
|
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Biochemistry,
37,
16192-16200.
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PDB code:
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W.R.Forsyth,
M.K.Gilson,
J.Antosiewicz,
O.R.Jaren,
and
A.D.Robertson
(1998).
Theoretical and experimental analysis of ionization equilibria in ovomucoid third domain.
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Biochemistry,
37,
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A.Motta,
P.Amodeo,
P.Fucile,
M.A.Castiglione Morelli,
B.Bremnes,
and
O.Bakke
(1997).
A new triple-stranded alpha-helical bundle in solution: the assembling of the cytosolic tail of MHC-associated invariant chain.
|
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Structure,
5,
1453-1464.
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B.A.Katz,
and
R.T.Cass
(1997).
In crystals of complexes of streptavidin with peptide ligands containing the HPQ sequence the pKa of the peptide histidine is less than 3.0.
|
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J Biol Chem,
272,
13220-13228.
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PDB codes:
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B.García-Moreno,
J.J.Dwyer,
A.G.Gittis,
E.E.Lattman,
D.S.Spencer,
and
W.E.Stites
(1997).
Experimental measurement of the effective dielectric in the hydrophobic core of a protein.
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| |
Biophys Chem,
64,
211-224.
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L.C.Ma,
and
S.Anderson
(1997).
Correlation between disulfide reduction and conformational unfolding in bovine pancreatic trypsin inhibitor.
|
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Biochemistry,
36,
3728-3736.
|
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C.M.Lukacs,
J.Q.Zhong,
M.I.Plotnick,
H.Rubin,
B.S.Cooperman,
and
D.W.Christianson
(1996).
Arginine substitutions in the hinge region of antichymotrypsin affect serpin beta-sheet rearrangement.
|
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Nat Struct Biol,
3,
888-893.
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PDB codes:
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J.Antosiewicz,
J.A.McCammon,
and
M.K.Gilson
(1996).
The determinants of pKas in proteins.
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| |
Biochemistry,
35,
7819-7833.
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M.P.Byrne,
C.A.Broomfield,
and
W.E.Stites
(1996).
Mustard gas crosslinking of proteins through preferential alkylation of cysteines.
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J Protein Chem,
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R.De Lorimier,
H.W.Hellinga,
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NMR studies of structure, hydrogen exchange, and main-chain dynamics in a disrupted-core mutant of thioredoxin.
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Protein Sci,
5,
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H.A.Nagarajaram,
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(1996).
A procedure for the prediction of temperature-sensitive mutants of a globular protein based solely on the amino acid sequence.
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Proc Natl Acad Sci U S A,
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Interleukin-6 (IL-6) antagonism by soluble IL-6 receptor alpha mutated in the predicted gp130-binding interface.
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J Biol Chem,
270,
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M.R.Ranjbar,
R.Wynn,
S.Anderson,
and
M.Laskowski
(1995).
Ionizable P1 residues in serine proteinase inhibitors undergo large pK shifts on complex formation.
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| |
J Biol Chem,
270,
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R.Wynn,
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F.M.Richards,
and
R.O.Fox
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Interactions in nonnative and truncated forms of staphylococcal nuclease as indicated by mutational free energy changes.
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| |
Protein Sci,
4,
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E.P.Baldwin,
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Core-packing constraints, hydrophobicity and protein design.
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| |
Curr Opin Biotechnol,
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F.J.Sigworth
(1994).
Voltage gating of ion channels.
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| |
Q Rev Biophys,
27,
1.
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Z.S.Hendsch,
and
B.Tidor
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Do salt bridges stabilize proteins? A continuum electrostatic analysis.
|
| |
Protein Sci,
3,
211-226.
|
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D.J.Weber,
E.H.Serpersu,
A.G.Gittis,
E.E.Lattman,
and
A.S.Mildvan
(1993).
NMR docking of the competitive inhibitor thymidine 3',5'-diphosphate into the X-ray structure of staphylococcal nuclease.
|
| |
Proteins,
17,
20-35.
|
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F.M.Richards,
and
W.A.Lim
(1993).
An analysis of packing in the protein folding problem.
|
| |
Q Rev Biophys,
26,
423-498.
|
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M.K.Gilson
(1993).
Multiple-site titration and molecular modeling: two rapid methods for computing energies and forces for ionizable groups in proteins.
|
| |
Proteins,
15,
266-282.
|
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|
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C.Eigenbrot,
and
A.A.Kossiakoff
(1992).
Structural consequences of mutation.
|
| |
Curr Opin Biotechnol,
3,
333-337.
|
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D.Shortle
(1992).
Mutational studies of protein structures and their stabilities.
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| |
Q Rev Biophys,
25,
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The most recent references are shown first.
Citation data come partly from CiteXplore and partly
from an automated harvesting procedure. Note that this is likely to be
only a partial list as not all journals are covered by
either method. However, we are continually building up the citation data
so more and more references will be included with time.
Where a reference describes a PDB structure, the PDB
code is
shown on the right.
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Links
