Biology Reference
In-Depth Information
biological role of a protein. Therefore, protein models can be readily
applied to interpret the impact mutations can have on the overall struc-
ture and, thus, function of a protein.
67,68
While objective scoring func-
tions to assess this impact are still not reliable, visual inspection associated
with a good knowledge and understanding of the rules underlying pro-
tein structure has proven useful in defining the broad reasons for mutant
malfunction (for concrete examples, see Refs. 67, 68 and 82). There is
an increasingly large body of data on naturally occurring mutations (over
43 000 human sequence variants are reported in SWISS-PROT) and
SNPs, of which a sizable proportion will alter the translated protein
sequences. Interpreting the potential functional effects of these mutants
will be crucial to elucidate the molecular basis of human diseases.
5.1.2. Planning site-directed mutagenesis experiments
One definite advantage of 3D structure and models in functional protein
analysis is that they provide a solid base for site-directed mutatgenesis
experiments aimed at the elucidation of the molecular function of pro-
teins. Even medium- and low-accuracy models can be used as solid
frameworks for experiment planning, and guide the selection of key
mutants designed to test functional hypotheses
83,84
(Fig. 1) or to modu-
late biophysical properties.
85
These experimentally generated mutants
complement the naturally occurring ones mentioned in the previous sub-
section and, together with the mapping of other facts such as glycosyla-
tion sites, greatly contribute to the elucidation of protein function.
1
For
instance, the comparative models generated for the Fas ligand, its protein
family members,
86
and its receptor illustrate how models can be applied
to (a) the understanding of the impact of naturally occurring muta-
tions
87-89
; (b) experimental mutagenesis; and (c) to the interpretation and
mapping of other known features, such as glycosylations, to understand
the finer molecular function of a protein.
5.2. Molecular Replacement
Solving the phase problem, i.e. knowing the phase of the diffracted waves,
in crystallography experiments is a crucial step towards reconstructing
