muramidase and cadmium-sulfide

A simple, rapid, straightforward and washing/separation free of in-solution digestion method for microwave-assisted tryptic digestion of proteins (cytochrome c, lysozyme and myoglobin) using bare TiO(2) nanoparticles (NPs) prepared in aqueous solution to serve as multifunctional nanoprobes in electrospray ionization mass spectrometry (ESI-MS) was demonstrated. The current approach is termed as 'on particle ionization/enrichment (OPIE)' and it can be applied in ESI-MS, atmospheric pressure-matrix-assisted laser desorption/ionization mass spectrometry (AP-MALDI-MS) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The bare TiO(2) NPs can assist, accelerate and effectively enhance the digestion efficiency, sequence coverage and detection sensitivity of peptides for the microwave-assisted tryptic digestion of proteins in ESI-MS. The reason is attributed to the fact that proteins or partially digested proteins are easily attracted or concentrated onto the surface of TiO(2) NPs, resulting in higher efficiency of digestion reactions in the microwave experiments. Besides, the TiO(2) NPs could act as a microwave absorber to accelerate and enrich the protein fragments in a short period of time (40-60 s) from the microwave experiments in ESI-MS. Furthermore, the bare TiO(2) NPs prepared in aqueous solution exhibit high adsorption capability toward the protein fragments (peptides); thus, the OPIE approach for detecting the digested protein fragments via ESI and MALDI ionization could be achieved. The current technique is also a washing and separation-free technique for accelerating and enriching microwave-assisted tryptic digestion of proteins in the ESI-MS and MALDI-MS. It exhibits potential to be widely applied to biotechnology and proteome research in the near future.

Topics: Animals; Cadmium Compounds; Cattle; Chickens; Cytochromes c; Hot Temperature; Microwaves; Molecular Probes; Muramidase; Nanoparticles; Peptide Fragments; Quantum Dots; Spectrometry, Mass, Electrospray Ionization; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Sulfides; Titanium; Trypsin

CdS quantum dots (∼ 5 nm) are used as multifunctional nanoprobes as an effective matrix for large proteins, peptides and as affinity probes for the enrichment of tryptic digest proteins (lysozyme, myoglobin and cytochrome c) in laser desorption/ionization time-of-flight mass spectrometry (LDI-TOF MS). The use of CdS quantum dots (CdS QDs) as the matrix allows acquisition of high resolution LDI mass spectra for large proteins (5000-80,000 Da). The enhancement of mass resolution is especially notable for large proteins such as BSA, HSA and transferrin (34-49 times) when compared with those obtained by using SA as the matrix. This technique demonstrates the potentiality of LDI-TOF-MS as an appropriate analytical tool for the analysis of high-molecular-weight biomolecules with high mass resolution. In addition, CdS QDs are also used as matrices for background-free detection of small biomolecules (peptides) and as affinity probes for the enrichment of tryptic digest proteins in LDI-TOF-MS.

Topics: Animals; Cadmium Compounds; Cytochromes c; Humans; Molecular Weight; Muramidase; Myoglobin; Peptides; Proteins; Quantum Dots; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Sulfides

For the first time, we utilized multifunctional nanoparticles composite (NPs composite) for matrix-assisted laser desorption/ionization mass spectrometric (MALDI-MS) analysis of peptides and proteins. Multiwalled carbon nanotubes doped with Cd(2+) ions and modified with cadmium sulfide NPs were synthesized by a chemical reduction method at room temperature. The multifunctional NPs composite applied for the analysis of peptides and microwave-digested proteins in the atmospheric pressure matrix-assisted laser desorption/ionization ion-trap and MALDI time-of-flight (TOF) mass spectrometry (MS) was successfully demonstrated. The maximum detection sensitivity for peptides in MALDI-MS was achieved by the adsorption of negatively charged peptides onto the surfaces of NP composite through electrostatic interactions. The optimal conditions of peptide mixtures were obtained at 20 min of incubation time using 1 mg of NPs composite when the pH of the sample solution was kept higher than the pI values of peptides. The potentiality of the NP composite in the preconcentration of peptides was compared with that of the individual NP by calculating the preconcentration factors (PF) and found that the NPs composite showed a 4-6 times of PF than the other NPs. In addition, the NPs composite was also applied as heat-absorbing materials for efficient microwave tryptic digestion of cytochrome c and lysozyme from milk protein in MALDI-TOF-MS analysis. We believe that the use of NPs composite technique would be an efficient and powerful preconcentrating tool for MALDI-MS for the study of proteome research.

Topics: Animals; Cadmium Compounds; Cytochromes c; Humans; Hydrogen-Ion Concentration; Manganese; Microwaves; Milk; Muramidase; Nanocomposites; Nanoparticles; Nanotubes, Carbon; Peptide Fragments; Proteins; Sensitivity and Specificity; Sodium Chloride; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Sulfides; Trypsin; Ubiquitin; Zinc Compounds