Multicellular organisms live, by and large, harmoniously with microbes. The cornea of the eye of an animal is almost always free of signs of infection. The insect flourishes without lymphocytes or antibodies. A plant seed germinates successfully in the midst of soil microbes. How is this accomplished? Both animals and plants possess potent, broad-spectrum antimicrobial peptides, which they use to fend off a wide range of microbes, including bacteria, fungi, viruses and protozoa. What sorts of molecules are they? How are they employed by animals in their defence? As our need for new antibiotics becomes more pressing, could we design anti-infective drugs based on the design principles these molecules teach us?

/pdf/antimicrobial-peptides-of-multicellular-organisms-24vvoqtunk.pdf

Antimicrobial peptides of multicellular organisms

https://www.cell.com/cell/pdf/S0092-8674(00)80172-5.pdf

The Dorsoventral Regulatory Gene Cassette spätzle/Toll/cactus Controls the Potent Antifungal Response in Drosophila Adults

A family of peptides with broad-spectrum antimicrobial activity has been isolated from the skin of the African clawed frog Xenopus laevis. It consists of two closely related peptides that are each 23 amino acids and differ by two substitutions. These peptides are water soluble, nonhemolytic at their effective antimicrobial concentrations, and potentially amphiphilic. At low concentrations they inhibit growth of numerous species of bacteria and fungi and induce osmotic lysis of protozoa. The sequence of a partial cDNA of the precursor reveals that both peptides derive from a common larger protein. These peptides appear to represent a previously unrecognized class of vertebrate antimicrobial activities.

https://www.pnas.org/content/pnas/84/15/5449.full.pdf

Magainins, a class of antimicrobial peptides from Xenopus skin: isolation, characterization of two active forms, and partial cDNA sequence of a precursor

Agents that increase the permeability of the outer membrane.

beta-Peptides: from structure to function.

Pig small intestine was used as starting material for a batchwise isolation of a peptide fraction enriched in antibacterial activities against Escherichia coli (anti-Ec factor) and against Bacillus megaterium (anti-Bm factor). Separation and further purification were by different types of chromatography. Sequence analysis showed the anti-Bm factor to be apparently similar to vasoactive intestinal peptide. The anti-Ec factor was found to have a 31-residue sequence that was cecropin-like. It was named cecropin P1 and its structure was confirmed by solid-phase synthesis. Synthetic cecropin P1 with and without C-terminal amide was assayed on eight different bacteria. Mobility comparison between synthetic and natural cecropin P1 indicates that the natural peptide has a free C-terminal carboxyl group.

https://www.pnas.org/content/pnas/86/23/9159.full.pdf

Antibacterial peptides from pig intestine: isolation of a mammalian cecropin

We recently isolated from pig intestine and characterized a 31-residue antibacterial peptide named cecropin P1 with activity against Escherichia coli and several other Gram-negative bacteria. The isolation involved a number of batch-wise steps followed by several chromatography steps. The continued investigation of these antibacterial peptides has now yielded another antibacterial peptide with high activity against both E. coli and Bacillus megaterium. Amino acid analysis showed a very high content of proline (49 mol%) and arginine (26 mol%), an intermediate level of phenylalanine and low levels of leucine, tyrosine, isoleucine, and glycine. The primary structure was determined by a combination of Edman degradation, plasma desorption mass spectrometry and C-terminal sequence analysis by carboxypeptidase Y degradation using capillary zone electrophoresis for detection of liberated residues. The calculated molecular mass was 4719.7 Da, which is in excellent agreement with 4719 Da obtained by plasma desorption mass spectrometry. The peptide was named PR-39 (proline-arginine-rich with a size of 39 residues). The lethal concentration of the peptide was determined against six Gram-negative and four Gram-positive strains of bacteria.

Amino acid sequence of PR-39. Isolation from pig intestine of a new member of the family of proline-arginine-rich antibacterial peptides.

All higher organisms can in some way defend or protect themselves against their natural flora of bacteria This applies also to insects which have highly effective immune systems, both cellular and humoral Many insects respond to an injection of live, non-pathogenic bacteria with the production of a potent cell-free antibacterial activity (for a review see Boman and Hultmark 1987) This phenomenon was first analysed at the molecular level using as a model system the Cecropia moth, Hyalophora cecropia (hereafter referred to as Cecropia) When a diapausing pupa is immunized it turns on the genes for immunity while the rest of the animal remains in a dormant state Immunized pupae of Cecropia are therefore a system for biological enrichment of the RNA and the proteins which are synthesized from the genes for immunity We have taken advantage of this fact both in the purification of 15 inducible immune proteins and in the isolation of immune RNA, used for the preparation of a cDNA library After a short period of RNA synthesis, the insects respond to live bacteria by the production of a potent antibacterial activity which is due to the synthesis of 15–20 immune proteins

https://febs.onlinelibrary.wiley.com/doi/epdf/10.1111/j.1432-1033.1991.tb16252.x

Cell-free immunity in Cecropia

Two cDNA clones containing coding information for cecropin B from the Cecropia moth (Hyalophora cecropia) were identified by means of a synthetic probe. Sequencing of the two inserts showed that cecropin B is processed from a 62-amino acid residue precursor molecule including a 26-residue leader peptide and a COOH-terminal glycine residue. The latter presumably donates the nitrogen of the amide group present on the COOH-terminal leucine residue of the mature cecropin B. The sequence deduced for the mature cecropin B differed in the COOH-terminal region from the tentative structure previously determined by carboxypeptidase digestion. To settle the discrepancy, cecropin B was synthesized according to the cDNA sequence with an amidated COOH-terminal leucine. Natural and synthetic cecropin B were found to be indistinguishable with respect to electrophoretic mobility and antibacterial activity against seven different bacteria. The COOH-terminal tetrapeptides were isolated from both natural and synthetic cecropin B and found to be indistinguishable. The correct sequence for cecropin B is (formula; see text).

https://www.pnas.org/content/pnas/82/8/2240.full.pdf

Molecular cloning, cDNA sequencing, and chemical synthesis of cecropin B from Hyalophora cecropia

To study the regulation of the immune genes in insects, we have cloned and sequenced the attacin gene locus of the giant silk moth Hyalophora cecropia The locus contains one acidic and one basic attacin gene as well as two pseudogenes, which are remnants of basic attacin genes A small insertion element was found within the locus The two functional attacin genes are transcribed in opposite directions and have two introns inserted at homologous positions A common sequence, GGGGATTCCT, is found at nucleotide position –48 in the acidic gene and at nucleotide position –58 in the basic gene Interestingly, this decanucleotide is similar to the consensus of the NF-kB-binding site Expression studies revealed that both attacins are strongly induced by phorbol 12-myristate 13-acetate, lipolysaccharide and bacteria However, only the acidic attacin gene showed a clear response to injury

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1991.tb15811.x

Jong-Youn Lee

Papers

Antibacterial peptides from pig intestine: isolation of a mammalian cecropin

Amino acid sequence of PR-39. Isolation from pig intestine of a new member of the family of proline-arginine-rich antibacterial peptides.

Cell-free immunity in Cecropia

Molecular cloning, cDNA sequencing, and chemical synthesis of cecropin B from Hyalophora cecropia

Structure and expression of the attacin genes in Hyalophora cecropia