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PDBsum entry 1f21
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* Residue conservation analysis
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Enzyme class:
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E.C.3.1.26.4
- ribonuclease H.
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Reaction:
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Endonucleolytic cleavage to 5'-phosphomonoester.
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DOI no:
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Protein Sci
9:1914-1921
(2000)
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PubMed id:
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Divalent metal cofactor binding in the kinetic folding trajectory of Escherichia coli ribonuclease HI.
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E.R.Goedken,
J.L.Keck,
J.M.Berger,
S.Marqusee.
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ABSTRACT
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Proteins often require cofactors to perform their biological functions and must
fold in the presence of their cognate ligands. Using circular dichroism
spectroscopy. we investigated the effects of divalent metal binding upon the
folding pathway of Escherichia coli RNase HI. This enzyme binds divalent metal
in its active site, which is proximal to the folding core of RNase HI as defined
by hydrogen/deuterium exchange studies. Metal binding increases the apparent
stability of native RNase HI chiefly by reducing the unfolding rate. As with the
apo-form of the protein, refolding from high denaturant concentrations in the
presence of Mg2+ follows three-state kinetics: formation of a rapid burst phase
followed by measurable single exponential kinetics. Therefore, the overall
folding pathway of RNase HI is minimally perturbed by the presence of metal
ions. Our results indicate that the metal cofactor enters the active site pocket
only after the enzyme reaches its native fold, and therefore, divalent metal
binding stabilizes the protein by decreasing its unfolding rate. Furthermore,
the binding of the cofactor is dependent upon a carboxylate critical for
activity (Asp10). A mutation in this residue (D10A) alters the folding kinetics
in the absence of metal ions such that they are similar to those observed for
the unaltered enzyme in the presence of metal.
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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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K.Ratcliff,
and
S.Marqusee
(2010).
Identification of residual structure in the unfolded state of ribonuclease H1 from the moderately thermophilic Chlorobium tepidum: comparison with thermophilic and mesophilic homologues.
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Biochemistry,
49,
5167-5175.
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H.Zhang,
T.Zhang,
K.Chen,
S.Shen,
J.Ruan,
and
L.Kurgan
(2009).
On the relation between residue flexibility and local solvent accessibility in proteins.
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Proteins,
76,
617-636.
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K.Ratcliff,
J.Corn,
and
S.Marqusee
(2009).
Structure, stability, and folding of ribonuclease H1 from the moderately thermophilic Chlorobium tepidum: comparison with thermophilic and mesophilic homologues.
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Biochemistry,
48,
5890-5898.
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PDB code:
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N.A.Bushmarina,
C.E.Blanchet,
G.Vernier,
and
V.Forge
(2006).
Cofactor effects on the protein folding reaction: acceleration of alpha-lactalbumin refolding by metal ions.
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Protein Sci,
15,
659-671.
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A.Schlessinger,
and
B.Rost
(2005).
Protein flexibility and rigidity predicted from sequence.
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Proteins,
61,
115-126.
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K.Klumpp,
J.Q.Hang,
S.Rajendran,
Y.Yang,
A.Derosier,
P.Wong Kai In,
H.Overton,
K.E.Parkes,
N.Cammack,
and
J.A.Martin
(2003).
Two-metal ion mechanism of RNA cleavage by HIV RNase H and mechanism-based design of selective HIV RNase H inhibitors.
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Nucleic Acids Res,
31,
6852-6859.
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S.Griffin,
C.L.Higgins,
T.Soulimane,
and
P.Wittung-Stafshede
(2003).
High thermal and chemical stability of Thermus thermophilus seven-iron ferredoxin. Linear clusters form at high pH on polypeptide unfolding.
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Eur J Biochem,
270,
4736-4743.
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S.Robic,
M.Guzman-Casado,
J.M.Sanchez-Ruiz,
and
S.Marqusee
(2003).
Role of residual structure in the unfolded state of a thermophilic protein.
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Proc Natl Acad Sci U S A,
100,
11345-11349.
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D.Apiyo,
and
P.Wittung-Stafshede
(2002).
Presence of the cofactor speeds up folding of Desulfovibrio desulfuricans flavodoxin.
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Protein Sci,
11,
1129-1135.
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G.Spudich,
S.Lorenz,
and
S.Marqusee
(2002).
Propagation of a single destabilizing mutation throughout the Escherichia coli ribonuclease HI native state.
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Protein Sci,
11,
522-528.
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PDB code:
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H.X.Zhou
(2002).
Toward the physical basis of thermophilic proteins: linking of enriched polar interactions and reduced heat capacity of unfolding.
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Biophys J,
83,
3126-3133.
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S.Robic,
J.M.Berger,
and
S.Marqusee
(2002).
Contributions of folding cores to the thermostabilities of two ribonucleases H.
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Protein Sci,
11,
381-389.
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PDB code:
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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
