Biology Reference
In-Depth Information
Fpg proteins is the composition of
the void-filling,
intercalation triad.
In EcoNei, all three residues are located on the same
5 loop and are
consecutive, that is, Gln69, Leu70, and Tyr71. In bacterial Fpg proteins,
NEIL1, MvNei1, and AthFpg, as mentioned earlier, the residues that consti-
tute the triad reside in two different loops. Another difference between
EcoFpg and the eukaryotic family members lies in the lesion-recognition
loop located in the C-terminal domain of both proteins. In bacterial Fpg, the
damaged base is everted from the DNA helix and is enveloped in a deep
pocket, which is capped by the
b
4/
b
9 loop at one end ( Fig. 3 ). This loop is
missing in EcoNei and the eukaryotic members for which a structure exists,
including NEIL1, AthFpg, and Neil3, and appears to be required for excising
8-oxoG. The vast repertoire of substrates of the Fpg/Nei family members and
their different preferences for opposite bases and DNA substrates (single-
stranded, double-stranded, or bubble DNA substrates) warrants further struc-
tural and biochemical scrutiny.
a
F/
b
IV. Glycosylases Search for Lesions
It has long been a question in the field as to how DNA glycosylases locate
the lesions they recognize in a sea of undamaged bases. This issue is compli-
cated by the fact that a glycosylase flips out the damage from the DNA helix
into its active-site pocket in order to perform its enzymatic function. Further-
more, glycosylases do not use biochemical energy and rely on thermal energy
so that lesions are found through random collisions between the glycosylase
and the DNA molecule. Because of this, three-dimensional diffusion is con-
sidered to be too inefficient to account for the number of lesions the glycosylase
must excise. Glycosylases are thought to bind to a nonspecific site on the DNA
molecule and slide along the DNA by one-dimensional diffusion until the
enzyme finds the lesion or disassociates from DNA. There have been a number
of hypotheses proposed for the lesion search itself. One model suggests that the
glycosylase binds to an extruded DNA lesion and then moves along the DNA,
testing every single base. 94 This appears to be unlikely as both kinetics 95 and
single-molecule studies 96,97 have shown that glycosylases scan DNA close to
diffusion limits, making it thermodynamically impossible for them to sequen-
tially extrude and examine every base. In the second model, the DNA glycosy-
lase traps a randomly extruded damaged base. This extrusion is more likely with
lesions as hydrogen bonding and stacking interactions would be altered com-
pared to the normal bases. This appears to be the mechanism used by uracil
DNA glycosylase. 98 In the third model, glycosylases slide along the DNA
molecule and are able to recognize their particular substrate by specific
Search WWH ::




Custom Search