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Figure 1.
Fig. 1. Identification of fully active disaggregated
mutants of hMIP-1  . A, the
number of single amino acid substitutions generated at each
residue in hMIP-1 . For a
full description of all substitutions see Craig et al. (41). The
number of alternative amino acid substitutions at each residue,
which expressed well (  20% hMIP-1
), is shown
above the origin and those which expressed poorly (  20% hMIP-1
) below the
origin. Cysteine residues at amino acid positions 10, 11, 34,
and 50 were not mutated to retain structural integrity. B and
subsequent panels refer to amino acid residues at which the
described properties have been identified are shown with a tall
histogram and variants which were assayed for a property but did
not meet the criteria (e.g. they were not disaggregated or they
were less potent) are shown by short histograms. B, variants
that expressed well and were disaggregated according to native
polyacrylamide gel electrophoresis. Disaggregated variants
migrated substantially further into the gel than hMIP-1 , which
remained near the well. C, variants that were disaggregated
according to sedimentation equilibrium AUC analysis.
Disaggregated variants possessed weight average molecular
weights 100,000 Da.
D, disaggregated variants that retained full competitive
receptor binding activity on FDCP-mix A4 cells. Fully active
variants were defined as those with IC[50] values 8 nM in this
cellular assay.
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