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1.
Appl Biochem Biotechnol ; 150(1): 97-111, 2008 Jul.
Article in English | MEDLINE | ID: mdl-18568300

ABSTRACT

A lectin-like protein from the seeds of Acacia farnesiana was isolated from the albumin fraction, characterized, and sequenced by tandem mass spectrometry. The albumin fraction was extracted with 0.5 M NaCl, and the lectin-like protein of A. farnesiana (AFAL) was purified by ion-exchange chromatography (Mono-Q) followed by chromatofocusing. AFAL agglutinated rabbit erythrocytes and did not agglutinate human ABO erythrocytes either native or treated with proteolytic enzymes. In sodium dodecyl sulfate gel electrophoresis under reducing and nonreducing conditions, AFAL separated into two bands with a subunit molecular mass of 35 and 50 kDa. The homogeneity of purified protein was confirmed by chromatofocusing with a pI = 4.0 +/- 0.5. Molecular exclusion chromatography confirmed time-dependent oligomerization in AFAL, in accordance with mass spectrometry analysis, which confers an alteration in AFAL affinity for chitin. The protein sequence was obtained by a liquid chromatography quadrupole time-of-flight experiment and showed that AFAL has 68% and 63% sequence similarity with lectins of Phaseolus vulgaris and Dolichos biflorus, respectively.


Subject(s)
Acacia/chemistry , Plant Lectins/isolation & purification , Seeds/chemistry , Amino Acid Sequence , Chitin/chemistry , Chromatography, Affinity , Chromatography, Gel , Chromatography, Ion Exchange , Fabaceae , Mass Spectrometry , Molecular Sequence Data , Molecular Weight , Plant Lectins/analysis , Plant Lectins/chemistry , Sequence Alignment , Sequence Analysis, Protein , Tandem Mass Spectrometry
2.
J Appl Microbiol ; 103(4): 1001-6, 2007 Oct.
Article in English | MEDLINE | ID: mdl-17897204

ABSTRACT

AIMS: The initial colonization of the tooth by streptococci involves their attachment to adsorbed components of the acquired pellicle. Avoiding this adhesion may be successful in preventing caries at early stages. Salivary mucins are glycoproteins that when absorbed onto hydroxyapatite may provide binding sites for certain bacteria. Algal lectins may be especially interesting for oral antiadhesion trials because of their great stability and high specificity for mucins. This work aimed to evaluate the potential of two algal lectins to inhibit the adherence of five streptococci species to the acquired pellicle in vitro. METHODS AND RESULTS: The lectins used were extracted from Bryothamnion triquetrum (BTL) and Bryothamnion seaforthii (BSL). Fluorescence microscopy was applied to visualize the ability of fluorescein isothiocyanate-labelled lectins to attach to the pellicle and revealed a similar capability for both lectins. Streptococcal adherence assays were performed using saliva-coated microtitre plates. BSL inhibited more than 75% of Streptococcus sanguis, Streptococcus mitis, Streptococcus sobrinus and Streptococcus mutans adherence, achieving 92% to the latter. BTL only obtained statistically significant results on S. mitis and S. sobrinus, whose adherence was decreased by 32.5% and 54.4%, respectively. CONCLUSION: Algal lectins are able to inhibit streptococcal adherence. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results support the proposed application of lectins in antiadhesion therapeutics.


Subject(s)
Bacterial Adhesion/drug effects , Dental Pellicle/microbiology , Lectins/pharmacology , Streptococcus/drug effects , Adsorption , Biofilms/growth & development , Durapatite/metabolism , Eukaryota/chemistry , Humans , Saliva/metabolism , Streptococcus/classification , Streptococcus/growth & development , Streptococcus/physiology
3.
Biochem Cell Biol ; 84(1): 49-54, 2006 Feb.
Article in English | MEDLINE | ID: mdl-16462889

ABSTRACT

The biochemical characterization of a new lectin (Hypnea cervicornis agglutinin or HCA) isolated from the Brazilian red alga H. cervicornis is reported. The haemagglutinating activity of the lectin was only inhibited by the glycoprotein porcine stomach mucin at a minimum inhibitory concentration of 19 microg x mL(-1). No haemagglutination inhibition was detected after the addition of simple sugars. The MALDI-TOF molecular masses of native and reduced and carbamidomethylated HCA were, respectively, 9196.6 Da and 9988.2 Da, indicating that the primary structure of the protein is crosslinked by 7 disulfide bonds. This unusual structural feature among lectins, along with its N-terminal sequence and amino-acid composition, clearly shows that HCA belongs to a protein family distinct from the isolectins Hypnin A1 and A2 isolated from the related Japanese alga Hypnea japonica. On the other hand, HCA displayed a high degree of similarity to the agglutinin from the Brazilian species Hypnea musciformis. Our data indicate the occurrence of structural diversity among lectins of closely related species living in distant ecosystems, i.e., the Pacific coast of Japan and the Atlantic coast of Brazil, and support the hypothesis that the lectin content (lectinome) might serve as a biomarker for taxonomical purposes.


Subject(s)
Agglutinins/chemistry , Agglutinins/isolation & purification , Rhodophyta/chemistry , Amino Acid Sequence , Amino Acids , Animals , Chromatography, Ion Exchange , Hemagglutination , Hemagglutination Tests , Molecular Sequence Data
4.
J Pharm Pharmacol ; 57(3): 375-81, 2005 Mar.
Article in English | MEDLINE | ID: mdl-15807994

ABSTRACT

PAL is a glucose/mannose-specific lectin isolated from Pisum arvense seeds. Previously, we demonstrated the capacity of other leguminous lectins to induce oedema formation and neutrophil stimulation. To investigate the potential pro-inflammatory activity of PAL, we have studied its ability to induce neutrophil migration into peritoneal cavities of rats and neutrophil chemotaxis in-vitro. The role of resident cells and sugar residues on PAL activity was analysed. PAL or saline (control) were administered intraperitoneally to rats, and total and differential leucocyte (macrophages, neutrophils and mast cells) counts were performed. The role of resident cells on the PAL effect was evaluated using three strategies: reducing the total resident cell population by lavage of rat cavities with saline; increasing macrophage population by treating animals with thioglycolate; and depleting mast cell population by subchronic treatment of rats with compound 48/80. PAL induced in-vitro and in-vivo neutrophil migration. In-vivo, PAL (50, 100, 200 and 300 microg) significantly (P < 0.05) and dose-dependently increased neutrophil migration by 600, 740, 900 and 940%, respectively, showing maximal effect 4 h after injection. PAL induced mononuclear cell migration. The neutrophil stimulatory effect of PAL was potentiated in animals treated with both thioglycolate and compound 48/ 80. The indirect lectin chemotactic effect was shown in rats injected with supernatant from cultured macrophages stimulated by PAL. In conclusion, PAL was shown to exhibit in-vivo and in-vitro proinflammatory activity. The in-vivo effect seemed to occur by a dual mechanism that was independent, but also dependent, on resident cells.


Subject(s)
Chemotaxis, Leukocyte , Neutrophils/drug effects , Plant Lectins/pharmacology , Seeds/chemistry , Animals , Dose-Response Relationship, Drug , Female , In Vitro Techniques , Macrophages, Peritoneal/metabolism , Male , Mast Cells/metabolism , Neutrophils/physiology , Peritoneal Cavity/cytology , Peritoneal Lavage , Rats , Rats, Wistar
5.
Protein Pept Lett ; 9(2): 159-66, 2002 Apr.
Article in English | MEDLINE | ID: mdl-12141914

ABSTRACT

A lectin from the red marine alga Hypnea musciformis (HML) was purified by extraction with 20 mM PBS, precipitation with 70% saturated ammonium sulphate, ion-exchange DEAE-Cellulose chromatography and RP-HPLC. The 9.3 kDa polypeptide agglutinates erythrocytes from various sources and shows oligomerization tendencies under certain MALDI-TOF/MS conditions. Preliminary N-terminal sequencing and biological assays strongly suggest that the HML may belong to a new class of algae lectins.


Subject(s)
Lectins/chemistry , Lectins/isolation & purification , Rhodophyta/metabolism , Animals , Chromatography, High Pressure Liquid , Chromatography, Ion Exchange , Dimerization , Humans , Protein Structure, Tertiary , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
6.
Eur J Biochem ; 268(16): 4414-22, 2001 Aug.
Article in English | MEDLINE | ID: mdl-11502201

ABSTRACT

A mannose/glucose-specific lectin was isolated from seeds of Parkia platycephala, the most primitive subfamily of Leguminosae plants. The molecular mass of the purified lectin determined by mass spectrometry was 47 946 +/- 6 Da (by electrospray ionization) and 47 951 +/- 9 Da (by matrix-assisted laser-desoption ionization). The apparent molecular mass of the lectin in solutions of pH in the range 4.5-8.5 determined by analytical ultracentrifugation equilibrium sedimentation was 94 +/- 3 kDa, showing that the protein behaved as a non-pH-dependent dimer. The amino-acid sequence of the Parkia lectin was determined by Edman degradation of overlapping peptides. This is the first report of the primary structure of a Mimosoideae lectin. The protein contained a blocked N-terminus and a single, nonglycosylated polypeptide chain composed of three tandemly arranged homologous domains. Each of these domains shares sequence similarity with jacalin-related lectin monomers from Asteraceae, Convolvulaceae, Moraceae, Musaceae, Gramineae, and Fagaceae plant families. Based on this homology, we predict that each Parkia lectin repeat may display a beta prism fold similar to that observed in the crystal structure of the lectin from Helianthus tuberosus. The P. platycephala lectin also shows sequence similarity with stress- and pathogen-upregulated defence genes of a number of different plants, suggesting a common ancestry for jacalin-related lectins and inducible defence proteins.


Subject(s)
Fabaceae/chemistry , Lectins/chemistry , Plants, Medicinal , Amino Acid Sequence , Molecular Sequence Data , Plant Lectins , Protein Structure, Quaternary , Repetitive Sequences, Amino Acid , Sequence Homology, Amino Acid , X-Ray Diffraction
7.
Cell Mol Life Sci ; 57(2): 343-50, 2000 Feb.
Article in English | MEDLINE | ID: mdl-10766029

ABSTRACT

The primary structure of a lectin isolated from the red alga Bryothamnion triquetrum was established by combination of Edman degradation of sets of overlapping peptides and mass spectrometry. It contains 91 amino acids and two disulphide bonds. The primary structure of the B. triquetrum lectin does not show amino acid sequence similarity with known plant and animal lectin structures. Hence, this protein may be the paradigm of a novel lectin family.


Subject(s)
Algal Proteins , Lectins/chemistry , Lectins/isolation & purification , Rhodophyta/chemistry , Amino Acid Sequence , Chromatography, High Pressure Liquid , Cysteine/analysis , Disulfides/analysis , Galactosamine/analysis , Glucosamine/analysis , Lectins/classification , Lectins/metabolism , Molecular Sequence Data , Molecular Weight , Peptide Fragments/chemistry , Peptide Fragments/isolation & purification , Peptide Fragments/metabolism , Protein Isoforms/chemistry , Protein Isoforms/isolation & purification , Protein Isoforms/metabolism , Sequence Analysis, Protein , Sequence Homology, Amino Acid , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
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