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pubmed:abstractText |
The repressor proteins of the LacI/GalR family exhibit significant similarity in their secondary and tertiary structures despite less than 35% identity in their primary sequences. Furthermore, the core domains of these oligomeric repressors, which mediate dimerization, are homologous with the monomeric periplasmic binding proteins, extending the issue of plasticity to quaternary structure. To elucidate the determinants of assembly, a structure-based alignment has been created for three repressors and four periplasmic binding proteins. Contact maps have also been constructed for the three repressor interfaces to distinguish any conserved interactions. These analyses show few strict requirements for assembly of the core N-subdomain interface. The interfaces of repressor core C-subdomains are well conserved at the structural level, and their primary sequences differ significantly from the monomeric periplasmic binding proteins at positions equivalent to LacI 281 and 282. However, previous biochemical and phenotypic analyses indicate that LacI tolerates many mutations at 281. Mutations at LacI 282 were shown to abrogate assembly, but for Y282D this could be compensated by a second-site mutation in the core N-subdomain at K84 to L or A. Using the link between LacI assembly and function, we have further identified 22 second-site mutations that compensate the Y282D dimerization defect in vivo. The sites of these mutations fall into several structural regions, each of which may influence assembly by a different mechanism. Thus, the 360-amino acid scaffold of LacI allows plasticity of its quaternary structure. The periplasmic binding proteins may require only minimal changes to facilitate oligomerization similar to the repressor proteins.
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