Surface salt bridges modulate the DNA site size of bacterial histone-like HU proteins.
Bacterial histone-like DNA-binding proteins are best known for their role in compacting the genomic DNA. Of these proteins, HU is ubiquitous and highly conserved across the eubacterial kingdom. Using the HBsu (Bacillus subtilis-encoded HU homologue) as a model, we explore here the molecular basis for the ability of some HU homologues to engage a longer approx. 35 bp DNA site as opposed to the much shorter sites reported for other homologues. Using electrophoretic mobility-shift assays, we show that the DNA site size for HBsu is approx. 10-13 bp and that a specific surface salt bridge limits the DNA site size for HBsu. Surface exposure of the highly conserved Lys3, achieved by substitution of its salt-bridging partner Asp26 with Ala, leads to enhanced DNA compaction by HBsu-D26A (where D26A stands for the mutant Asp26-->Ala), consistent with the interaction of Lys3 with the ends of a 25 bp duplex. Both HBsu and HBsu-D26A bend DNA, as demonstrated by their equivalent ability to promote ligase-mediated DNA cyclization, indicating that residues involved in mediating DNA kinks are unaltered in the mutant protein. We suggest that Lys3 is important for DNA wrapping due to its position at a distance from the DNA kinks where it can exert optimal leverage on flanking DNA and that participation of Lys3 in a surface salt bridge competes for its interaction with DNA phosphates, thereby reducing the occluded site size.
Kamau
Edwin
E
Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA.
Tsihlis
Nick D
ND
Simmons
L Alice
LA
Grove
Anne
A
D019943:Amino Acid Substitution; D001412:Bacillus subtilis / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry*; D004269:DNA, Bacterial / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry*; D001411:Geobacillus stearothermophilus / Q000737:chemistry; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011487:Protein Conformation
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; C040330:histone-like protein HU, bacteria