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VOL. 73 (3), 703-746, 2007 ARE ADRENOMEDULLIN POSITIVE MODULATORS NOVEL...

The Lamarckian Genetic Algorithm (LGA) was used as the search
engine. The active site was defined using AutoGrid. The grid size was
set to 128 x 60 x 60 points with a grid spacing of 0.375 Å centered
on the catalytic zinc of each 1hov model. Step sizes of 2 Å for
translation and 60º for rotation were chosen. The maximum number
of energy evaluations was set to 250000. For each of the 100
independent runs, a maximum number of 27000 LGA operations
were generated on a single population of 100 individuals. The
compounds were studied with no restraints between the ZBG and
the zinc atom. Other variables were set by using AutoDock Tools
(ADT) (26).

Perl-based Filter

A perl program was developed in order to classify the compounds
into three different categories depending on the coordination with
the zinc ion: trigonal bypiramide coordination, square-based
pyramide coordination and no coordinating compound using the
mean atom distance and angles displacement published by Alberts
and colleagues (Figure 3) (28).

Biological Assays

The MMP-2 Colorimetric Drug Discovery Kit (purchased from
Biomol International, Lp., Exeter, UK) was selected for the biological
evaluation of MMP-2 inhibition by the selected compounds 1n, 2m
and 2n. Inhibition assays were carried out in duplicate at four
different concentrations (100 µM, 25 µM, 6.5 µM and 3.25 µM). The
inhibitor NNGH (29) was also included as a prototypic control
inhibitor. A commercially available negative control, with reported
no MMP inhibitory activity, was treated similarly (30).

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