Molecular recognition: structural and energetic aspects of preorganization, substrate specificity, and oligomerization
Abstract
Understanding the structural and energetic characteristics of the interactions
between biomolecules is a center of focus in the world of drug design. The approach
taken here is to analyze three fairly different molecular recognition phenomenons. First,
the energetics of binding by a Shc based peptide to monomeric and domain-swapped
dimeric Grb2-SH2 is analyzed. The peptide bound to monomer with over 15-fold higher
affinity than to domain-swapped dimeric Grb2-SH2. Two crystal structures were solved
which suggest the molecular basis for the observed binding thermodynamic differences
are alternate conformations of the EF loop in monomer and domain-swapped dimer
Grb2-SH2. Secondly, the impact of introducing a cyclopropane ring, to act as a
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conformational constraint, into Shc based peptides that bind the Grb2-SH2 domain is
studied. Isothermal titration calorimetry experiments showed the constrained ligands
bound with higher affinity compared to flexible controls. Interestingly, the higher
affinity for the constrained compounds was the result of an enthalpic advantaged, in
contrast to an expected entropic advantage for the restriction of rotors. Several crystal
structures were solved, which suggest the conformationally constrained ligands likely
bound as a more rigid complex compared to the flexible control. Lastly, the structural
basis for the substrate specificity profiles of three PC-PLCBc protein variants is
investigated. The crystal structures of these protein variants were solved, and structures
of complexes between a phosphatidylserine analogue and wild-type PC-PLCBc and two
phosphatidylserine specific protein variants were also obtained. Those structures suggest
active site water molecules and the side chain conformations of residues at the 4th and
55th positions play a crucial role in the substrate specificity profile of the protein variants
studied.
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