Peptide profile of human acquired enamel pellicle using MALDI tandem MS

Vitorino, Rui; Calheiros-Lobo, Maria João; Duarte, José Alberto; Domingues, Pedro; Amado, Francisco

doi:10.1002/jssc.200700486

Cited by 43 publications

(52 citation statements)

References 43 publications

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“…However, in agreement with Gibbins et al (2014), blotting experiments showed that statherin adsorbed preferentially on more hydrophobic substrata. The more substantial statherin adsorption on hydrophobized silica surfaces is consistent with previous reports (Santos et al 2008;Vitorino et al 2008). This is explained by the prevalence of the hydrophobic interaction developed between the hydrophobic substrata and the C-terminal part of the protein which also has a hydrophobic character.…”

Section: Discussionsupporting

confidence: 91%

Influence of substratum hydrophobicity on salivary pellicles: organization or composition?

Aroonsang¹,

Sotres²,

El-Schich³

et al. 2014

Biofouling

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Different physico-chemical properties (eg adsorption kinetics, thickness, viscoelasticity, and mechanical stability) of adsorbed salivary pellicles depend on different factors, including the properties (eg charge, roughness, wettability, and surface chemistry) of the substratum. Whether these differences in the physico-chemical properties are a result of differences in the composition or in the organization of the pellicles is not known. In this work, the influence of substratum wettability on the composition of the pellicle was studied. For this purpose, pellicles eluted from substrata of different but well-characterized wettabilities were examined by means of sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The results showed that substratum hydrophobicity did not have a major impact on pellicle composition. In all substrata, the major pellicle components were found to be cystatins, amylases and large glycoproteins, presumably mucins. In turn, interpretation of previously reported data based on the present results suggests that variations in substratum wettability mostly affect the organization of the pellicle components.

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Section: Discussionsupporting

confidence: 91%

Influence of substratum hydrophobicity on salivary pellicles: organization or composition?

Aroonsang¹,

Sotres²,

El-Schich³

et al. 2014

Biofouling

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“…The resulting peptides were extracted with trifluoroacetic acid/acetonitrile/water, and then analyzed by MALDI-TOF, using sinapinic acid as matrix on a micro MX linear and reflection TOF mass spectrometer (Waters, Milford, MA). Screening of the LC-MALDI plate was performed in the MS positive reflector mode, using 1200 laser shots at a collision energy of 2 kV with air as collision gas at a pressure of 2.610 À7 Torr (Vitorino et al, 2008). The spectra were processed and analyzed by the Global Protein Server Workstation (Applied Biosystems, Foster City, CA), which uses internal MASCOT software (Matrix Science, Boston, MA) for searching the peptide mass fingerprints and MS/MS data.…”

Section: Maldi-tof/tof-ms Analysismentioning

confidence: 99%

Bio-sintering processes in hexactinellid sponges: Fusion of bio-silica in giant basal spicules from Monorhaphis chuni☆

Müller¹,

Wang²,

Burghard³

et al. 2009

Journal of Structural Biology

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“…In addition, several of the ions arose from unknown proteins. Using our proposed methodology of on-plate digestion more than 100 different tryptic peptides were characterised (12). This led to the identification of 12 distinct salivary proteins.…”

mentioning

confidence: 99%

On-plate digestion using a commercial microfraction collector for nano-HPLC matrix-assisted laser desorption/ionization tandem time-of-flight protein analysis

Vitorino

Guedes

Tomer

et al. 2008

Analytical Biochemistry

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A new method for on-plate protein digestion and MALDI MS analysis is proposed involving an automated one-step sample separation using nanoflow-HPLC followed by nanoliter fraction collection and on-plate digestion with trypsin. This procedure uses a commercial automatic nanoliter fraction collection system for on-line spotting of the eluent onto a MALDI target. After protein digestion, the reaction is stopped by the addition of acidified matrix using the same automated system. Collected spots are subsequently analyzed using a MALDI-TOF/TOF mass spectrometer for protein sequencing and identification. Keywordsacquired enamel pellicle; LC-MS/MS; on-plate digestion; proteomic; MALDI-TOF/TOF Gel-free methods have been developed based on liquid separations coupled on-line with mass spectrometry (LC-MS) for proteomic analyses of complex biological systems (1);(2)). The sequence information obtained from peptide fragment fingerprints and database searching is often sufficient for identification (1-3). However, one of the limitations of this approach is the difficulty in identification of protein isoforms or highly homologous proteins. One approach to circumvent this problem proposed by Zheng et al. (4), is based on protein separation using a capillary column followed by fraction collection on MALDI targets pre-coated with trypsin for on-plate digestion, followed by MALDI-TOF/TOF analysis.We have developed a new "hands-off" method for on-line protein digestion and MALDI analysis based on a robotic system coupled to a nano-HPLC. This system offers improved reproducibility and ease of handling for routine proteomics studies using standard gel-free mass spectrometry and tandem mass spectrometry screening procedures. In order to assess this approach, we performed on-plate plate digestion with standard proteins such as equine myoglobin (Sigma M0630), bovine insulin (Sigma, St. Louis, MO, I5500) and bovine serum albumin (Sigma L3908). Protein solutions were prepared at an initial concentration of 1mg/ mL and then diluted, so that 0.5 μL of the protein solution corresponded to 20 pmol, 100 pmol, 200 pmol, 600 pmol and 12 nmol of protein. Aliquots, 0.5 μL were spotted onto a MALDI target, and the spot was then incubated with 0.2 μL of a trypsin solution (0.2 μg/μL in 25mMCorrespondence, Rui M P Vitorino, Department of Chemistry, University of Aveiro, Aveiro, Portugal, rvitorino@ua. The database search parameters were: mass tolerance of 40 ppm for precursor ions and 0.3 Da for fragment ions; trypsin digestion with two missed cleavages; variable modifications such as N -terminal Gln, pyroGlutamic acid, methionine oxidation, N-terminal acetylation and phosphorylation of serine, threonine and tyrosine residues. A positive identification was accepted at the 95% confidence level.As expected, good protein coverage was achieved through the approach of in-solution digestion of the protein standards at all tested concentrations. Similar results were obtained using our protein on-plate digestion approach based on the number of detec...

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Peptide profile of human acquired enamel pellicle using MALDI tandem MS

Cited by 43 publications

References 43 publications

Influence of substratum hydrophobicity on salivary pellicles: organization or composition?

Influence of substratum hydrophobicity on salivary pellicles: organization or composition?

Bio-sintering processes in hexactinellid sponges: Fusion of bio-silica in giant basal spicules from Monorhaphis chuni☆

On-plate digestion using a commercial microfraction collector for nano-HPLC matrix-assisted laser desorption/ionization tandem time-of-flight protein analysis

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