Chemistry Reference
In-Depth Information
Figure 15.5
Alignment of metal ions in the two-metal-ion mechanism. Two metal ions,
preferably Mg
2+
, are jointly coordinated by at least one absolutely conserved Asp and the
scissile phosphate, which is the
a
-phosphate of (d)NTP in the case of polymerase. Metal
ion A is directly coordinated to the nucleophile and metal ion B to the leaving group. The
reaction products are 3
-PO
4
. (Reproduced from Yang, W.
et al
., Making and
breaking nucleic acids: two-Mg
2+
-ion catalysis and substrate specifi city,
Mol. Cell
, 2006,
22
,
5-13)
′
-OH and 5
′
the bases and bisected by the plane of the trigonal core of the pentacovalent inter-
mediate (Figure 15.5). Based on the particular coordination geometry of the two
metal ions, Steitz and Steitz proposed a generally applicable two-metal-ion mecha-
nism, by which metal ion A acts like a Lewis base to reduce the p
K
a of a nucleophile
and facilitate its deprotonation and metal ion B stabilizes the pentacovalent inter-
mediate and the 3
oxyanion leaving group.
84
In the last 14 years, many structures of polymerases and nucleases using
two - metal - ion catalysis have been determined.
1,29 - 31
In addition to facilitating
catalysis, the metal ions appear to play a critical role in enhancing substrate recogni-
tion and catalytic specifi city for both polymerases and type IIP REases. By being
sensitive to the coordination environment, the metal ions induce misalignment
between a nonpreferential substrate and catalytic residues, thus greatly reducing
the catalytic effi ciency. Catalysis occurs effi ciently only with a preferred substrate.
The enhancement of substrate recognition is not limited to the fi delity of DNA
and RNA polymerases, and sequence and methylation specifi c nucleases, but also
extends to structure-specifi c nucleases, e.g. RNase H and argonaute. Although the
latter nucleases have no explicit sequence specifi city, RNase H only cleaves the
RNA strand in a RNA/DNA hybrid and not in dsRNA, while argonaute functions
in tandem with a duplex ruler and generates cleavage products of a specifi c
length.
85
′
