Yu-Xi Gong

Structure–activity relationships of fowlicidin‐1, a cathelicidin antimicrobial peptide in chicken

Cationic antimicrobial peptides are naturally occurring antibiotics that are actively being explored as a new class of anti‐infective agents. We recently identified three cathelicidin antimicrobial peptides from chicken, which have potent and broad‐spectrum antibacterial activities in vitro (Xiao Y, Cai Y, Bommineni YR, Fernando SC, Prakash O, Gilliland SE & Zhang G (2006) J Biol Chem281, 2858–2867). Here we report that fowlicidin‐1 mainly adopts an α‐helical conformation with a slight kink induced by glycine close to the center, in addition to a short flexible unstructured region near the N terminus. To gain further insight into the structural requirements for function, a series of truncation and substitution mutants of fowlicidin‐1 were synthesized and tested separately for their antibacterial, cytolytic and lipopolysaccharide (LPS)‐binding activities. The short C‐terminal helical segment after the kink, consisting of a stretch of eight amino acids (residues 16–23), was shown to be critically involved in all three functions, suggesting that this region may be required for the peptide to interact with LPS and lipid membranes and to permeabilize both prokaryotic and eukaryotic cells. We also identified a second segment, comprising three amino acids (residues 5–7) in the N‐terminal flexible region, that participates in LPS binding and cytotoxicity but is less important in bacterial killing. The fowlicidin‐1 analog, with deletion of the second N‐terminal segment (residues 5–7), was found to retain substantial antibacterial potency with a significant reduction in cytotoxicity. Such a peptide analog may have considerable potential for development as an anti‐infective agent.

Activity and Structural Comparisons of Solution Associating and Monomeric Channel-Forming Peptides Derived from the Glycine Receptor M2 Segment

A number of channel-forming peptides derived from the second transmembrane (TM) segment (M2) of the glycine receptor alpha(1) subunit (M2GlyR), including the 22-residue sequence NK(4)-M2GlyR p22 wild type (WT) (KKKKPARVGLGITTVLTMTTQS), induce anion permeation across epithelial cell monolayers. In vitro assays suggest that this peptide or related sequences might function as a candidate for ion channel replacement therapy in treating channelopathies such as cystic fibrosis (CF). The wild-type sequence forms soluble associations in water that diminish its efficacy. Introduction of a single substitution S22W at the C-terminus, NK(4)-M2GlyR p22 S22W, eliminates the formation of higher molecular weight associations in solution. The S22W peptide also reduces the concentration of peptide required for half-maximal anion transport induced across Madin-Darby canine kidney cells (MDCK) monolayers. A combination of 2D double quantum filtered correlation spectroscopy (DQF-COSY), total correlation spectroscopy (TOCSY), nuclear Overhauser effect spectroscopy (NOESY), and rotating frame nuclear Overhauser effect spectroscopy (ROESY) data were recorded for both the associating WT and nonassociating S22W peptides and used to compare the primary structures and to assign the secondary structures. High-resolution structural studies were recorded in the solvent system (40% 2,2,2-Trifluoroethanol (TFE)/water), which gave the largest structural difference between the two peptides. Nuclear Overhauser effect crosspeak intensity provided interproton distances and the torsion angles were measured by spin-spin coupling constants. These constraints were put into the DYANA modeling program to generate a group of structures. These studies yielded energy-minimized structures for this mixed solvent environment. Structure for both peptides is confined to the 15-residue transmembrane segments. The energy-minimized structure for the WT peptide shows a partially helical extended structure. The S22W peptide adopts a bent conformation forming a hydrophobic pocket by hydrophobic interactions.

Solution structure, antibacterial activity, and expression profile of Manduca sexta moricin.

In response to wounding or infection, insects produce a battery of antimicrobial peptides (AMPs) and other defense molecules to kill the invading pathogens. To study their structures, functions, and transcriptional regulation, we synthesized Manduca sexta moricin, a 42‐residue peptide (GKIPVKAIKQAGKVIGKGLRAINIAGTTHDVVSFFRPKKKKH, 4539 Da). The compound exhibited potent antimicrobial activities against a broad spectrum of Gram‐positive and Gram‐negative bacteria with a minimum inhibitory concentration of 1.4 µM. The mRNA levels of M. sexta moricin increased substantially in fat body and hemocytes after the larvae were challenged with bacterial cells. We determined the solution structure of this AMP by two‐dimensional 1H1H ‐nuclear magnetic resonance spectroscopy. The tertiary structure is composed of an eight‐turn α‐helix spanning almost the entire peptide. Insights of relationships between the structure and function are also presented. Copyright

NMR Structure/Function Relationships of Peptides Corresponding to the C1B1 Region of PKCγ

The region (101-112) of C1B domain in PKC gamma plays a crucial role in the activation of the enzyme and subsequent gap junction inhibition. Substitution studies on peptides correlating to the C1B region show that a flexible structure and ability to be phosphorylated on serine 109 are critical for this purpose.