Order and Disorder in the Domain Organization of the Plasmid Partition Protein KorB

Abstract

The plasmid partition protein KorB has a dual role: it is essential for the correct segregation of the low copy number broad host range RK2 plasmid while also being an important regulator of transcription. KorB belongs to the ParB family of proteins, and partitioning in RK2 has been studied as a simplified model of bacterial chromosome segregation. Structural information on full-length ParB proteins is limited, mainly due to the inability to grow crystals suitable for diffraction studies. We show, using CD and NMR, that KorB has regions of significant intrinsic disorder and hence it adopts a multiplicity of conformations in solution. The biophysical data are consistent with bioinformatic predictions based on the amino acid sequence that the N-terminal region and also the region between the central DNA-binding domain and the C-terminal dimerization domain are intrinsically disordered. We have used small angle x-ray scattering data to determine the ensemble of solution conformations for KorB and selected deletion mutants, based on models of the known domain structures. This conformational range of KorB is likely to be biologically required for DNA partitioning and for binding to a diverse set of partner proteins.

The Evidence of Large-Scale DNA-Induced Compaction in the Mycobacterial Chromosomal ParB

“…The conformational flexibility and intrinsic disorder were previously reported for KorB, which is a plasmid-based functional analog of ParB. 35 KorB and tbParB share a somewhat similar pattern of predicted disordered regions. 25,35 Based on that mentioned above, we expected to find either global or local intrinsic disorder in the apo tbParB.…”

Characterization of Intrinsically Disordered Proteins by Analytical Ultracentrifugation

“…The observation of larger f/f 0 values for globular proteins is therefore attributed to asymmetry. Values of the frictional ration for intrinsically disordered proteins are relatively high: 2.8 for nucleoporin (Denning et al, 2003), 2.1 for securin (Sánchez-Puig et al, 2005), 1.9 for KorB (Rajasekar et al, 2010), and 1.6 for neuroligin (Paz et al, 2008). Whether these higher values of f/f 0 reflect asymmetry of a compact protein or a larger value of δ for an unstructured species cannot be determined by analytical ultracentrifugation.…”

“…In studies of intrinsically disordered proteins the value of the curve-fitting parameter also includes any additional spreading as the result of variation in s across the ensemble of species comprising a given oligomeric state. Of the various software packages available for numerical solution of the Lamm equation (BPCFIT, LAMM, SEDANAL, SEDFIT, SEDPHAT, SVEDBERG, ULTRASCAN), the SEDFIT program (Schuck, 1998(Schuck, , 2000 has been the procedure of choice for studies of intrinsically disordered proteins (Batra-Safferling et al, 2006;Paz et al, 2008;Rajasekar et al, 2010). Its application to the neurologin 3 distributions presented in Fig.…”

“…In studies of intrinsically disordered proteins, this solvated radius has routinely been obtained (Adkins & Lumb, 2002;Batra-Safferling et al, 2006;Denning et al, 2003;Kalthoff et al, 2002;Paz et al, 2008;Rajasekar et al, 2010;Sánchez-Puig et al, 2005) from size-exclusion chromatography experiments by comparing its elution characteristics with those for globular proteins with defined Stokes radii (Laurent & Killander, 1964;Siegel & Monty, 1966).…”

“…For some proteins, this nonrigid structure occurs throughout the entire molecule; but in others, e.g., the plasmid partition protein KorB (Rajasekar et al, 2010) there are intrinsically disordered regions amidst sections exhibiting the compact structure that is amenable to delineation by X-ray crystallography. Historically, intrinsically disordered proteins have been much ignored in the literature, as the drive has always been to isolate globular proteins for structural and functional studies.…”