Biology Reference
In-Depth Information
A. DNA Binding
One key difference between the endonuclease activities of
B. subtilis
and
N. gonorrhoeae
MutL is the requirement for the ATPase domain of the protein.
B. subtilis
MutL-CTD does not bind DNA on its own and, consequently, its
endonuclease activity is undetectable in the absence of the ATPase domain of
the protein.
96
Conversely, both the full-length and the C-terminal domain of
N.
gonorrhoeae
MutL bind DNA and have measurable endonuclease activity.
92
Accordingly, the dimerization domain of
N. gonorrhoeae
MutL can bind DNA
even in the absence of the ATPase domain of the protein,
92
reinforcing the idea
that the lack of activity in
B. subtilis
MutL-CTD is due to the low affinity of this
domain for DNA.
The N-terminal domains of
E. coli
, human, and yeast homologs of MutL bind
DNA even in the absence of the dimerization domain of the protein.
98,101,107
However, the dimerization domains of
E. coli
and human MutL
a
do not bind
DNA on their own (J. W. and A. G., unpublished results).
42
The endonuclease
activity of eukaryotic MutL
a
has not been assayed in the absence of the ATPase
domain and, hence, it is unclear whether this domain is strictly required for the
activity of MutL. The conformational change imposed by the association of
the two ATPase domains upon nucleotide binding should bring DNA bound at
the inner groove of the N-terminal domain of the protein in close proximity to the
endonuclease domain of the protein (
Fig. 2
).
100
Indeed, a recent study where
ATPase and endonuclease domains of
A. aeolicus
MutL were obtained separately
reveals that the ATPase domain stimulates the endonuclease activity in a Zn
2
รพ
-
dependent manner,
116
suggesting the direct interaction between the two domains
of the protein. This implicitly suggests that ATP binding may stimulate the
endonuclease activity of the protein by providing a mechanism to bypass the
DNA binding defect of the endonuclease domain.
96
Therefore, the lack of DNA
binding at the C-terminal domains of human MutL
a
and
B. subtilis
MutL may
entail a powerful mechanism to avoid indiscriminate nicking of DNA.
B. Stimulatory Effect of the Processivity Clamp
PCNA and RFC (the clamp loader for PCNA) stimulate the endonuclease
activity of human and yeast MutL
a
.
67,94
This stimulation has been attributed to
the direct interaction between MutL
a
and PCNA.
139
The interaction between
these two proteins has been demonstrated and a conserved motif located in the
C terminus of yMLH1 has been deemed important for PCNA binding.
73,76
However, while the consensus sequence of this motif (
Q
IX[
L/I
]XX
F
A) loosely
resembles a PCNA-interacting motif (
Q
XX[
M/L/I
]XX
F
[Y/F]), its conformation
is markedly different from other bona fide PCNA-interacting motifs. The N
terminus of PCNA-interacting motifs is usually extended and typically makes a
short antiparallel
b
-zipper with the C-terminal
b
-strand of the clamp, while the
