Proteolytic activity of bovine lactoferrin

Massucci, Maria; Giansanti, Francesco; Nino, Giovanna Di; Turacchio, Manola; Giardi, Maria Federica; Botti, Dario; Ippoliti, Rodolfo; Giulio, Barbara De; Siciliano, Rosa Anna; Donnarumma, Giovanna; Valenti, Piera; Bocedi, Alessio; Polticelli, Fabio; Ascenzi, Paolo; Antonini, Giovanni

doi:10.1023/b:biom.0000027700.90780.45

Cited by 25 publications

(22 citation statements)

References 21 publications

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“…1c) was reported for mast cells alpha-chymase in conjunction with the attenuation of allergic response [55]. Two cleavage sites at the edges of the β-strands were reported for lactoferrin (Fig.5a) as a result of autoproteolytic activity of this iron binding protein associated with mammalian non-immune defense against pathogens [56]. Cleavage of an internal strand of a β-sheet, which, at the same time, is the N-terminus of the protein, was registered for actin and two different types of proteases (Fig.…”

Section: Discussionmentioning

confidence: 94%

Structural Determinants of Limited Proteolysis

et al. 2011

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Limited or regulatory proteolysis plays a critical role in many important biological pathways like blood coagulation, cell proliferation, and apoptosis. A better understanding of mechanisms that control this process is required for discovering new proteolytic events and for developing inhibitors with potential therapeutic value. Two features that determine the susceptibility of peptide bonds to proteolysis are the sequence in the vicinity of the scissile bond and the structural context in which the bond is displayed. In this study we assessed statistical significance and predictive power of individual structural descriptors and combination thereof for the identification of cleavage sites. The analysis was performed on a dataset of >200 proteolytic events documented in CutDB for a variety of mammalian regulatory proteases and their physiological substrates with known 3D structures. The results confirmed the significance and provided a ranking within three main categories of structural features: exposure > flexibility > local interactions. Among secondary structure elements, the largest frequency of proteolytic cleavage was confirmed for loops and lower but significant frequency for helices. Limited proteolysis has lower albeit appreciable frequency of occurrence in certain types of β-strands, which is in contrast with some previous reports. Descriptors deduced directly from the amino acid sequence displayed only marginal predictive capabilities. Homology-based structural models showed a predictive performance comparable to protein substrates with experimentally established structures. Overall, this study provided a foundation for accurate automated prediction of segments of protein structure susceptible to proteolytic processing and, potentially, other post-translational modifications.

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Section: Discussionmentioning

confidence: 94%

Structural Determinants of Limited Proteolysis

et al. 2011

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“…N-␣-Benzyloxycarbonyl-Phe-Arg-7-amido-4-methylcoumarin (Z-Phe-Arg-AMC) (Sigma) was used as a substrate, as described previously (42), and Lf-catalyzed hydrolysis was determined in PBS at room temperature (21°C). Lf was used at a concentration of 10 M, and the substrate concentration ranged from 1 to 200 M. The cleavage of the substrate was followed fluorometrically at 460 nm with an excitation wavelength of 355 nm by monitoring the increase with a Victor microtiter plate reader (Wallac).…”

Section: Methodsmentioning

confidence: 99%

“…The serine protease activity of aLf has been described extensively previously (33,42,52,57,68). To test whether this activity was responsible for S. aureus killing, aliquots of bovine aLf and human aLf were treated with PMSF, a potent inhibitor of serine proteases, and used in killing assays.…”

Section: Isda Is An Lf Binding Proteinmentioning

confidence: 99%

“…The serine protease activity resides in the N lobe, but the precise location is a matter of contention. One study proposed that it consists of a serine-lysine catalytic dyad located in the area adjacent to the interlobe cleft of human Lf (33), but in another study, pK a shift calculations indicated that several serine residues of bovine Lf display the high nucleophilicity required to potentially catalyze substrate cleavage (42). Definitive identification of the active site awaits further study.…”

Section: Vol 76 2008mentioning

confidence: 99%

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IsdA ProtectsStaphylococcus aureusagainst the Bactericidal Protease Activity of Apolactoferrin

Clarke

Foster

2008

Infect Immun

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An important facet of the Staphylococcus aureus host-pathogen interaction is the ability of the invading bacterium to evade host innate defenses, particularly the cocktail of host antimicrobial peptides. In this work, we showed that IsdA, a surface protein of S. aureus which is required for nasal colonization, binds to lactoferrin, the most abundant antistaphylococcal polypeptide in human nasal secretions. The presence of IsdA on the surface of S. aureus confers resistance to killing by lactoferrin. In addition, the bactericidal activity of lactoferrin was inhibited by addition of phenylmethylsulfonyl fluoride, implicating the serine protease activity of lactoferrin in the killing of S. aureus. Recombinant IsdA was a competitive inhibitor of lactoferrin protease activity. Reciprocally, antibody reactive to IsdA enhanced killing of S. aureus. Thus, IsdA can protect S. aureus against lactoferrin and acts as a protease inhibitor.

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“…They also found that Lf blocks Enteropathogenic E. coli adherence, hemolysis and induction of actin polymerisation in HEp2 cells as a result of the Lf-mediated degradation of E. coli secreted proteins A, B and D (EspABD) [63,64]. Massucci et al [65] characterised in vitro the proteolytic activity of bovine Lf towards synthetic substrates. The substrate specifi city is similar to that of trypsin, and serine protease inhibitors inhibit this catalytic activity, although the values of the catalytic parameters (k cat , K m ) are several orders of magnitude lower than those of trypsin.…”

Section: Antibacterial Activity Related To Proteolysismentioning

confidence: 99%

Lactoferrin

Valenti

Antonini

2005

Cell. Mol. Life Sci.

408

165

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The first function attributed to lactoferrin (Lf), an iron binding protein belonging to the non-immune natural defences, was antimicrobial activity that depended on its capacity to sequester iron. Iron-independent microbicidal activities, requiring direct interaction between this cationic protein and microbial surface components, were later demonstrated. Many other anti-microbial and anti-viral functions have since been ascribed to Lf. In mucosal secretions, iron and Lf modulate the motility and aggregation of pathogenic bacteria. Lf inhibits bacterial adhesion on abiotic surfaces through ionic binding to biomaterials, or specific binding to bacterial structures or both. Lf inhibition of bacterial adhesion to host cells requires Lf binding to bacteria and/or host cells. Lf hinders microbial internalization by binding to both glycosaminoglycans and bacterial proteins which can be degraded by Lf-mediated proteolysis. Moreover, Lf internalisation and localisation to the host cell nuclei could modulate bacterial entry into cells through gene regulation. Finally, the capability of Lf to exert antiviral activity, through its binding to host cells and/or viral particles, strengthens the idea that it is an important brick in the mucosal wall, effective against both microbial and viral attacks.

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Proteolytic activity of bovine lactoferrin

Cited by 25 publications

References 21 publications

Structural Determinants of Limited Proteolysis

Structural Determinants of Limited Proteolysis

IsdA ProtectsStaphylococcus aureusagainst the Bactericidal Protease Activity of Apolactoferrin

Lactoferrin

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