Inhibition of a plant virus infection by analogs of melittin.

An approach that enables identification of specific synthetic peptide inhibitors of plant viral infection is reported. Synthetic analogs of melittin that have sequence and structural similarities to an essential domain of tobacco mosaic virus coat protein were found to possess highly specific antiviral activity. This approach involves modification of residues located at positions analogous to those that are critical for virus assembly. The degree of inhibition found correlates well with sequence similarities between the viral capsid protein and the melittin analogs studied as well as with the induced conformational changes that result upon interaction of the peptides and ribonucleic acid.

Retro and retroenantio analogs of cecropin-melittin hybrids.

Hybrid analogs of cecropin A (CA) and melittin (M), which are potent antibacterial peptides, have been synthesized. To understand the structural requirements for this antibacterial activity, we have also synthesized the enantio, retro, and retroenantio isomers of two of the hybrids and their N-terminally acetylated derivatives. All analogs of CA(1-13)M(1-13)-NH2 were as active as the parent peptide against five test bacterial strains, but one bacterial strain was resistant to the retro and retroenantio derivatives. Similarly, all analogs of CA(1-7)M(2-9)-NH2 were active against four strains, while two strains were resistant to the retro and retroenantio analogs containing free NH3+ end groups, but acetylation restored activity against one of them. From these data it was concluded that chirality of the peptide was not a critical feature, and full activity could be achieved with peptides containing either all L- or all D-amino acids in their respective right-handed or left-handed helical conformations. For most of the bacterial strains, the sequence of these peptides or the direction of the peptide bonds could be critical but not both at the same time. For some strains, both needed to be conserved.

Automated de novo sequencing of proteins by tandem high-resolution mass spectrometry

A de novo sequencing program for proteins is described that uses tandem MS data from electron capture dissociation and collisionally activated dissociation of electrosprayed protein ions. Computer automation is used to convert the fragment ion mass values derived from these spectra into the most probable protein sequence, without distinguishing Leu/Ile. Minimum human input is necessary for the data reduction and interpretation. No extra chemistry is necessary to distinguish N- and C-terminal fragments in the mass spectra, as this is determined from the electron capture dissociation data. With parts-per-million mass accuracy (now available by using higher field Fourier transform MS instruments), the complete sequences of ubiquitin (8.6 kDa) and melittin (2.8 kDa) were predicted correctly by the program. The data available also provided 91% of the cytochrome c (12.4 kDa) sequence (essentially complete except for the tandem MS-resistant region K13–V20 that contains the cyclic heme). Uncorrected mass values from a 6-T instrument still gave 86% of the sequence for ubiquitin, except for distinguishing Gln/Lys. Extensive sequencing of larger proteins should be possible by applying the algorithm to pieces of ≈10-kDa size, such as products of limited proteolysis.