Antibacterial proteins

Smith, V. J., Fernandes, J. M. O., Kemp, G. D., and Flauston, C. (2008). Crustins Enigmatic WAP domain-containing antibacterial proteins from crustaceans. Dev. Comp. Immunol. 32, 758-772. 

James, S., Holmstrom, C., and Kjelleberg, S., Purification and characterization of a novel antibacterial protein from the marine bacterium D2, Appl. Environ. Microbiol., 62, 2783, 1996. 

Oncology, NADH-ubiquinone red. Antibacterial, inflammation Antibacterial, protein biosynthesis... 

Patynowski et al. (2002) showed that yeasts produce an unidentified inhibitory factor (maybe a toxic metabolite) that could be responsible for the inhibition of bacterial growth. These results could explain the antagonism between yeasts and malolactic bacteria, since yeasts are known to produce compounds during alcoholic fermentation such as ethanol, SO2, medium-chain fatty acids and antibacterial proteins/peptides (Weeks et al. 1969 De Oliva et al. 2004 Comitini et al. 2005 Osborne and Edwards 2007). The nature and quantity of peptides and other molecules released by yeasts are different depending on winemaking techniques and the yeast strain. 

Wibowo et al (1988) proposed that S. cerevisiae may inhibit O. oeni through the production of antibacterial proteins/peptides and not through the production of S02 and ethanol. In support, Dick et al (1992) isolated two proteins produced by S. cerevisiae, which showed activity against O. oeni Despite these findings, very few further studies investigating the production of antibacterial proteins/peptides by wine yeast have been undertaken. 

In addition to S02 and antibacterial proteins/peptides, medium-chain fatty acids produced by yeast during alcoholic fermentation have also been implicated in the inhibition of malolactic bacteria (Carrete et al, 2002 Edwards and Beelman, 1987 Lonvaud-Funel et al, 1985). Inhibition of Saccharomyces species and some LAB by medium-chain fatty acids has been reported in grape juice and silage (Pederson et al, 1961 Woolford, 1975). Although this hypothesis has not been conclusively shown, Lonvaud-Funel et al (1985) and... 

Malec et al. (M2) discovered the synthesis of antibacterial proteins and nuclear RNA in isolated leukocyte nuclei. In granulocytes, more than one half of mRNA is unstable, quickly and continuously renewing itself and ensuring protein synthesis (Cl). 

M2. Malec, J., Karnacka, L., and Wajnorowska, M., Protein and nucleic acid synthesis in isolated leucocyte nuclei and some evidence for the production of an antibacterial protein. Exp. Cell Res. 34, 188-191 (1964). 

Fly blood does not normally contain substances that kill bacteria, but flies inoculated with bacteria rapidly accumulate antibacterial proteins (ABs) in their blood. Wild type Drosophila have at least three different antibacterial proteins based on isoelectric points. Genetic variants identify structural genes for these antibacterial proteins. A DNA sequence that can encode a conserved portion of moth and fleshfly antibacterial proteins has been used to synthesize a complementary oligonucleotide probe. This probe recognizes a messenger RNA that appears in the fat body of Drosophila and Medflies only after they have been inoculated with bacteria. Bacteria-sensitive lethal mutations were induced to identify genes necessary for flies to survive a bacterial infection. 

This communication reports studies on the humoral antibacterial response in Drosophila and Ceratitis capitata, the Mediterranean fruit fly. The goal of our work is to understand the molecular mechanisms that protect flies from bacterial infection. Here we examine three questions 1) What genes encode antibacterial proteins 2) Does the antibacterial response involve the accumulation of new messenger RNAs And 3), what genes are necessary to survive a bacterial attack Answers to these questions will help us to determine the potential of blocking the immune system to control populations of insect pests. 

Genes Specifying Antibacterial Protein Structure. Adult flies and larvae of Drosophila and Ceratitis do not normally contain in their blood proteins that specifically kill bacteria. However, when animals are inoculated with Enterobacter cloacae, potent antibacterial activity appears in the blood (11,12,17,18). Antibacterial activity is detected in the blood of adult Drosophila melanogaster flies within two hours after inoculation, and is still detectable sixty days later (11). Investigation of this activity by isoelectric focusing reveals several blood proteins with antibacterial activity... 

Stock At contains only AB8.7 and AB9.1, while stock Am has only AB7.1. The wild stock Oregon R has all three spots. Stock Cu lacks all three antibacterial proteins found in wild type but has a novel band at pI7.6. In addition, stock Cu is quite sensitive to infection. We are currently mapping these mutations to identify the chromosomal location of genes that affect AB structure. 

Figure 1. Different species of flies have different patterns of antibacterial activity after isoelectric focusing. Dro, Drosophila melanogaster Med, Ceratitis capitata Mel, Dacus cucurbitae Dor, Dacus dorsalis. Antibacterial proteins (ABs) are labeled according to their isoelectric points.

The results of these experiments 1) provide genetic variants for genes that control the structure of antibacterial proteins 2) show that specific RNAs accumulate after a bacterial infection and 3)... 

Figure 6. Bacteria-sensitive lethal mutants are inducible for antibacterial proteins.

The immune system of Drosophila seems to have two main subsystems, one involving the antibacterial proteins and the other identified by bacteria-sensitive lethal mutations. The humoral subsystem results in the production of diffusible antibacterial proteins of at least three species. Stock Cu, which lacks the ABs found in wild type Drosophila and which is correspondingly more sensitive to infection, blocks a part of this subsystem. That the ABs do help flies to survive bacterial infections was shown by inducing ABs to appear in bsl mutants by injection of dead bacteria and then finding that the inoculated mutants would survive an otherwise lethal injection of live bacteria. Evidently inoculation induces processes (perhaps secretion of ABs) that can overcome the deficiency in the bsl mutations. Similar experiments showed that inoculation protects locusts from a lethal dose of bacteria (19). 

Some of the ABs may have structural homology to antibacterial proteins isolated from the flesh fly (20,21) since an oligonucleotide probe that can encode a portion of the sarcotoxin protein recognizes an immune-specific RNA in Drosophila fat body cells, the cellular origin of Drosophila s ABs (unpubl.). The induction mechanism must work relatively rapidly since we found immune-specific RNA in fat body cells within 6 hours after inoculation. Because of its homology to sarcotoxin, we assume that the immune-specific transcript detected by the oligonucleotide probe encodes an antibacterial protein. 

Insects fight back against infecting bacteria by producing antibacterial proteins [ 105]. These include cecropins, attacins, defensins,lysozyme, diptericins, sarcotoxins, apidaecin, and abaecin. The molecules either cause lysis or are bacteriostatic, and also attack parasites. 

Although historically most useful antibiotics have come from spore forming microorganisms, marine organisms have yielded the candidate antitumor peptide didemnin B [77327-50-0] (2) and cytostatic peptides such as the patellamides (3). Many of the marine peptides have little or no antimicrobial activity. Antibacterial peptides called magainins are found in frog skin (4) and antibacterial proteins called defensins are found in mammalian white blood cells (5). The commercially important insecticidal proteins from Bacillus thuringensis (6) are not discussed herein nor are the numerous peptide siderophores (7,8), which, except for the albomycins (9), are usually not antimicrobial. 

Steiner, H., Hultmark, D., Engstrom, A., Ben-nich, H., Boman, H.G. Sequence and specificity of two antibacterial proteins involved in insect immunity. Nature 1981, 292, 246-248... 

Smith, V. J. Purification and characterization of a cysteine-rich 11. 5-kDa antibacterial protein from the granular haemocytes of the shore crab, Carcinus maenas. Eur. J. Biochem. 

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