Antimicrobial peptides have been isolated and characterized from tissues and organisms representing virtually every kingdom and phylum, ranging from prokaryotes to humans. Yet, recurrent structural and functional themes in mechanisms of action and resistance are observed among peptides of widely diverse source and composition. Biochemical distinctions among the peptides themselves, target versus host cells, and the microenvironments in which these counterparts convene, likely provide for varying degrees of selective toxicity among diverse antimicrobial peptide types. Moreover, many antimicrobial peptides employ sophisticated and dynamic mechanisms of action to effect rapid and potent activities consistent with their likely roles in antimicrobial host defense. In balance, successful microbial pathogens have evolved multifaceted and effective countermeasures to avoid exposure to and subvert mechanisms of antimicrobial peptides. A clearer recognition of these opposing themes will significantly advance our understanding of how antimicrobial peptides function in defense against infection. Furthermore, this understanding may provide new models and strategies for developing novel antimicrobial agents, that may also augment immunity, restore potency or amplify the mechanisms of conventional antibiotics, and minimize antimicrobial resistance mechanisms among pathogens. From these perspectives, the intention of this review is to illustrate the contemporary structural and functional themes among mechanisms of antimicrobial peptide action and resistance.

Mechanisms of Antimicrobial Peptide Action and Resistance

基礎講座 電気泳動(Electrophoresis)

Antimicrobial host defense peptides are produced by all complex organisms as well as some microbes and have diverse and complex antimicrobial activities. Collectively these peptides demonstrate a broad range of antiviral and antibacterial activities and modes of action, and it is important to distinguish between direct microbicidal and indirect activities against such pathogens. The structural requirements of peptides for antiviral and antibacterial activities are evaluated in light of the diverse set of primary and secondary structures described for host defense peptides. Peptides with antifungal and antiparasitic activities are discussed in less detail, although the broad-spectrum activities of such peptides indicate that they are important host defense molecules. Knowledge regarding the relationship between peptide structure and function as well as their mechanism of action is being applied in the design of antimicrobial peptide variants as potential novel therapeutic agents.

Peptide Antimicrobial Agents

An adequate supply of saliva is critical to the preservation and maintenance of oral tissue. Clinicians often do not value the many benefits of saliva until quantities are decreased. Much is written on the subject of salivary hypofunction, but little attention is paid to normal salivary flow and function. This article is a brief, up-to-date overview of the literature on the basics of normal salivary composition, flow, and function. A review of the literature was conducted using MEDLINE and Healthstar (1944 through 1999); articles were selected for inclusion on the basis of relevance and significance to the clinician.

A review of saliva: Normal composition, flow, and function

The potential of luminescent semiconductor quantum dots (QDs) to enable development of hybrid inorganic-bioreceptor sensing materials has remained largely unrealized. We report the design, formation and testing of QD‐protein assemblies that function as chemical sensors. In these assemblies, multiple copies of Escherichia coli maltose-binding protein (MBP) coordinate to each QD by a C-terminal oligohistidine segment and function as sugar receptors. Sensors are selfassembled in solution in a controllable manner. In one configuration, a β-cyclodextrin-QSY9 dark quencher conjugate bound in the MBP saccharide binding site results in fluorescence resonance energy-transfer (FRET) quenching of QD photoluminescence. Added maltose displaces the β-cyclodextrin-QSY9, and QD photoluminescence increases in a systematic manner. A second maltose sensor assembly consists of QDs coupled with Cy3-labelled MBP bound to β-cyclodextrin-Cy3.5. In this case, the QD donor drives sensor function through a two-step FRET mechanism that overcomes inherent QD donor‐acceptor distance limitations. Quantum dot‐biomolecule assemblies constructed using these methods may facilitate development of new hybrid sensing materials.

Self-assembled nanoscale biosensors based on quantum dot FRET donors

https://www.jbc.org/content/263/16/7472.full.pdf

Histatins, a novel family of histidine-rich proteins in human parotid secretion. Isolation, characterization, primary structure, and fungistatic effects on Candida albicans.

J. LI, E.J. HELMERHORST, C.W. LEONE, R.F. TROXLER, T. YASKELL, A.D. HAFFAJEE, S.S. SOCRANSKY AND F.G. OPPENHEIM. 2004. Aims: To elucidate the first colonizers within in vivo dental biofilm and to establish potential population shifts that occur during the early phases of biofilm formation. Methods and Results: AcheckerboardDNA-DNA hybridization assay was employed to identify 40 different bacterial strains. Dental biofilm samples were collected from 15 healthy subjects, 0, 2, 4 and 6 h after tooth cleaning and the composition of these samples was compared with that of whole saliva collected from the same individuals. The bacterial distribution in biofilm samples was distinct from that in saliva, confirming the selectivity of the adhesion process. In the very early stages, the predominant tooth colonizers were found to be Actinomyces species. The relative proportion of streptococci, in particular Streptococcus mitis and S. oralis, increased at the expense of Actinomyces species between 2 and 6 h while the absolute level of Actinomyces remained unaltered. Periodontal pathogens such as Tannerella forsythensis (Bacteroides forsythus), Porphyromonas gingivalis and Treponema denticola as well as Actinobacillus actinomycetemcomitans were present in extremely low levels at all the examined time intervals in this healthy group of subjects. Conclusion: The data provide a detailed insight into the bacterial population shifts occurring within the first few hours of biofilm formation and show that the early colonizers of the tooth surface predominantly consist of beneficial micro-organisms. Significance and Impact of the Study: The early colonizers of dental plaque are of great importance in the succession stages of biofilm formation and its overall effect on the oral health of the host.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1365-2672.2004.02420.x

Identification of early microbial colonizers in human dental biofilm

The acquired enamel pellicle is an organic film covering the surfaces of teeth. When this film was first discovered, it was thought to be of embryologic origin. Only in the middle of this century did it become clear that it was acquired after tooth eruption. Initially, the small amounts of material that could be obtained have virtually limited the investigation of pellicle proteins to amino acid analysis. Nevertheless, this technique revealed that the pellicle is mainly proteinaceous and is formed by selective adsorption of salivary proteins on tooth enamel. Later, immunologic techniques allowed for the identification of many salivary and fewer non-salivary proteins as constituents of pellicle. However, to this date, isolation and direct biochemical characterization of in vivo pellicle protein have not been possible, because only a few micrograms can be obtained from a single donor. Therefore, the composition and structure of the acquired enamel pellicle are still essentially unknown. Information on the f...

Saliva and dental pellicle--a review.

The proteome of whole saliva, in contrast to that of serum, is highly susceptible to a variety of physiological and biochemical processes. First, salivary protein secretion is under neurologic control, with protein output being dependent on the stimulus. Second, extensive salivary protein modifications occur in the oral environment, where a plethora of host- and bacteria-derived enzymes act on proteins emanating from the glandular ducts. Salivary protein biosynthesis starts with the transcription and translation of salivary protein genes in the glands, followed by post-translational processing involving protein glycosylation, phosphorylation, and proteolysis. This gives rise to salivary proteins occurring in families, consisting of structurally closely related family members. Once glandular secretions enter the non-sterile oral environment, proteins are subjected to additional and continuous protein modifications, leading to extensive proteolytic cleavage, partial deglycosylation, and protein-protein complex formation. All these protein modifications occur in a dynamic environment dictated by the continuous supply of newly synthesized proteins and removal by swallowing. Understanding the proteome of whole saliva in an environment of continuous turnover will be a prerequisite to gain insight into the physiological and pathological processes relevant to oral health, and be crucial for the identification of meaningful biomarkers for oral disease.

Saliva: a Dynamic Proteome

We have previously shown that histatins 1, 3, and 5 are homologous, histidine-rich proteins present in human parotid and submandibular secretions which contain 38, 32, and 24 amino acids, respectively. Interest in these proteins stems from the fact that histatins exhibit candidacidal and candidastatic activities. The goal of the present investigation was a detailed functional characterization of these anticandidal activities of histatins at the levels of killing of blastoconidia, killing of germinated cells, and inhibition of germination by using three bioassays. Candidacidal activities were evaluated at several ionic strengths, in the presence of different mono- and divalent ions, and at multiple pH values. In addition, the susceptibility of Candida albicans in different growth phases to histatins was investigated. While all three major human histatins demonstrated candidacidal activities, they differed in their abilities to kill blastoconidia and germinated cells, with histatin 5 being the most active, histatin 3 showing moderate activity, and histatin 1 exhibiting the lowest level of activity. For the inhibition of germination, however, histatin 3 exhibited more activity than either histatin 1 or histatin 5. The candidacidal activity of histatins was inversely proportional to both the ionic strength and the divalent cation concentration in the medium. Stepwise reduction of the pH of the assay medium enhanced the candidacidal activities of histatins 1 and 3, while the activity of histatin 5 was pH independent over the range of pHs 4 to 8. C. albicans in log-phase growth was more susceptible to histatins 1 and 3 than cells in stationary phase. Cells in either growth phase were still more vulnerable to histatin 5 than to histatins 1 and 3. The results obtained establish the functional relationship of the major histatins with respect to both their fungicidal and fungistatic activities and provide insights into their activities under ionic and pH conditions likely to be encountered in vivo in the oral cavity. Moreover, the data point towards possible mechanisms responsible for the anticandidal activities of histatins.

Frank G. Oppenheim

Papers

Histatins, a novel family of histidine-rich proteins in human parotid secretion. Isolation, characterization, primary structure, and fungistatic effects on Candida albicans.

Identification of early microbial colonizers in human dental biofilm

Saliva and dental pellicle--a review.

Saliva: a Dynamic Proteome

Anticandidal activity of major human salivary histatins.