Structural and Functional Diversity of Acidic Scorpion Potassium Channel Toxins

Background: Although the basic scorpion K + channel toxins (KTxs) are well-known pharmacological tools and potential drug candidates, characterization the acidic KTxs still has the great significance for their potential selectivity towards different K + channel subtypes. Unfortunately, research on the acidic KTxs has been ignored for several years and progressed slowly. Principal Findings: Here, we describe the identification of nine new acidic KTxs by cDNA cloning and bioinformatic analyses. Seven of these toxins belong to three new a-KTx subfamilies (a-KTx28, a-KTx29, and a-KTx30), and two are new members of the known k-KTx2 subfamily. ImKTx104 containing three disulfide bridges, the first member of the a-KTx28 subfamily, has a low sequence homology with other known KTxs, and its NMR structure suggests ImKTx104 adopts a modified cystine-stabilized a-helix-loop-b-sheet (CS-a/b) fold motif that has no apparent a-helixs and b-sheets, but still stabilized by three disulfide bridges. These newly described acidic KTxs exhibit differential pharmacological effects on potassium channels. Acidic scorpion toxin ImKTx104 was the first peptide inhibitor found to affect KCNQ1 channel, which is insensitive to the basic KTxs and is strongly associated with human cardiac abnormalities. ImKTx104 selectively inhibited KCNQ1 channel with a Kd of 11.69 mM, but was less effective against the basic KTxs-sensitive potassium channels. In addition to the ImKTx104 toxin, HeTx204 peptide, containing a cystine-stabilized a-helix-loop-helix (CS-a/a) fold scaffold motif

The Antibiotic Novobiocin Binds and Activates the ATPase That Powers Lipopolysaccharide Transport

Novobiocin is an orally active antibiotic that inhibits DNA gyrase by binding the ATP-binding site in the ATPase subunit. Although effective against Gram-positive pathogens, novobiocin has limited activity against Gram-negative organisms due to the presence of the lipopolysaccharide-containing outer membrane, which acts as a permeability barrier. Using a novobiocin-sensitive <i>Escherichia coli</i> strain with a leaky outer membrane, we identified a mutant with increased resistance to novobiocin. Unexpectedly, the mutation that increases novobiocin resistance was not found to alter gyrase, but the ATPase that powers lipopolysaccharide (LPS) transport. Co-crystal structures, biochemical, and genetic evidence show novobiocin directly binds this ATPase. Novobiocin does not bind the ATP binding site but rather the interface between the ATPase subunits and the transmembrane subunits of the LPS transporter. This interaction increases the activity of the LPS transporter, which in turn alters the permeability of the outer membrane. We propose that novobiocin will be a useful tool for understanding how ATP hydrolysis is coupled to LPS transport