No abstract

SummaryA central feature of broad host range IncP-1 plasmids is the set of regulatory circuits that tightly control plasmid core functions under steady-state conditions. Cooperativity between KorB and either KorA or TrbA repressor proteins is a key element of these circuits and deletion analysis has implicated the conserved C-terminal domain of KorA and TrbA in this interaction. By NMR we show that KorA and KorB interact directly and identify KorA amino acids that are affected on KorB binding. Studies on mutants showed that tyrosine 84 (or phenylalanine, in some alleles) is dispensable for repressor activity but critical for the specific interaction with KorB in both in vivo reporter gene assays and in vitro electrophoretic mobility shift and co-purification assays. This confirms that direct and specific protein-protein interactions are responsible for the cooperativity observed between KorB and its corepressors and lays the basis for determining the biological importance of this cooperativity.

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.

No abstract

Objectives: Corn silk is an underutilized part of corn which possesses great medicinal importance. The present study was planned to determine the phytochemical composition and antioxidant potential of corn silk extracts, obtained by different extraction methods using a series of solvents with increasing polarity. Methods: Three extraction methods 1) individual extraction in each solvent, 2) consecutive extraction in solvents of increasing polarity and 3) consecutive extraction of crude methanolic extract in solvents of increasing polarity and a series of five solvents with increasing polarity were used for extraction of phytochemicals. The extracts were analyzed for phytochemical composition and antioxidant potential. Results: Corn silk was found to contain a variety of bioactive phytochemical compounds including phenolic acids, flavonoids, ascorbic acid, tannins and cardiac glycosides. The corn silk phytochemicals were extracted more in high polarity solvents which showed comparatively good phytochemical composition and strong antioxidants potential. Regression analysis of experimental data showed a polarity dependent increase in extraction yield and phytochemical content and free radical scavenging capacity of extracts obtained by various extraction methods. Conclusion: The water extract obtained by individual extraction showed a comparatively high extraction yield and phytochemical content while that obtained by consecutive extraction of crude methanolic extract showed high ability to scavenge free radicals. The study advocates the corn silk as a good source of antioxidant phytochemicals and suggests the use of polar solvents and individual extraction method for extraction of corn silk phytochemicals.