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Differences in protein changes between stress-induced premature senescence and replicative senescence states

Aan GJ, Hairi HA, Makpol S, Rahman MA, Karsani SA

Electrophoresis, 2013 Aug;34(15):2209-17.
PMID: 23712505 DOI: 10.1002/elps.201300086

Replicative senescence and stress-induced premature senescence (SIPS) cells are known to share certain traits. However, whether these cells are different at the protein level is unclear. Thus, this study has utilized proteomics to identify differences in the proteomes of replicative senescence and SIPS cells compared to normal cells. Replicative senescence was induced by serial passage of normal cells in culture. SIPS was established by exposure to H2 O2 at a subcytotoxic concentration of 20 μM for two weeks. Following 2DE, protein profiles were compared and protein spots that changed in abundance were identified by MALDI-TOF MS. Quantitative real-time RT-PCR was then performed to evaluate the transcript expression of selected altered proteins. A total of 24 and 10 proteins were found to have changed in abundance in replicative senescence and SIPS cells, respectively, when compared to young cells. Quantitative RT-PCR revealed that nine genes showed the same direction of change as observed in the proteomics analysis. Very little overlap was observed between proteins that changed in replicative senescence and SIPS cells, suggesting that although both SIPS and replicative senescence cells share hallmarks of cellular senescence, they were different in terms of proteins that changed in abundance.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext Gamma-tocotrienol treatment increased peroxiredoxin-4 expression in HepG2 liver cancer cell line

Abdul Rahman Sazli F, Jubri Z, Abdul Rahman M, Karsani SA, Md Top AG, Wan Ngah WZ

BMC Complement Altern Med, 2015;15:64.
PMID: 25886747 DOI: 10.1186/s12906-015-0590-y

To determine the antiproliferative effect of gamma-tocotrienol (GTT) treatment on differential protein expression in HepG2 cells.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext Purification and Characterization of a Novel NAD(P)+-Farnesol Dehydrogenase from Polygonum minus Leaves

Ahmad-Sohdi NA, Seman-Kamarulzaman AF, Mohamed-Hussein ZA, Hassan M

PLoS One, 2015;10(11):e0143310.
PMID: 26600471 DOI: 10.1371/journal.pone.0143310

Juvenile hormones have attracted attention as safe and selective targets for the design and development of environmentally friendly and biorational insecticides. In the juvenile hormone III biosynthetic pathway, the enzyme farnesol dehydrogenase catalyzes the oxidation of farnesol to farnesal. In this study, farnesol dehydrogenase was extracted from Polygonum minus leaves and purified 204-fold to apparent homogeneity by ion-exchange chromatography using DEAE-Toyopearl, SP-Toyopearl, and Super-Q Toyopearl, followed by three successive purifications by gel filtration chromatography on a TSK-gel GS3000SW. The enzyme is a heterodimer comprised of subunits with molecular masses of 65 kDa and 70 kDa. The optimum temperature and pH were 35°C and pH 9.5, respectively. Activity was inhibited by sulfhydryl reagents, metal-chelating agents and heavy metal ions. The enzyme utilized both NAD+ and NADP+ as coenzymes with Km values of 0.74 mM and 40 mM, respectively. Trans, trans-farnesol was the preferred substrate for the P. minus farnesol dehydrogenase. Geometrical isomers of trans, trans-farnesol, cis, trans-farnesol and cis, cis-farnesol were also oxidized by the enzyme with lower activity. The Km values for trans, trans-farnesol, cis, trans-farnesol and cis, cis-farnesol appeared to be 0.17 mM, 0.33 mM and 0.42 mM, respectively. The amino acid sequences of 4 tryptic peptides of the enzyme were analyzed by MALDI-TOF/TOF-MS spectrometry, and showed no significant similarity to those of previously reported farnesol dehydrogenases. These results suggest that the purified enzyme is a novel NAD(P)+-dependent farnesol dehydrogenase. The purification and characterization established in the current study will serve as a basis to provide new information for recombinant production of the enzyme. Therefore, recombinant farnesol dehydrogenase may provide a useful molecular tool in manipulating juvenile hormone biosynthesis to generate transgenic plants for pest control.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext MALDI-target integrated platform for affinity-captured protein digestion

Ahmad-Tajudin A, Adler B, Ekström S, Marko-Varga G, Malm J, Lilja H, et al.

Anal Chim Acta, 2014 Jan 7;807:1-8.
PMID: 24356215 DOI: 10.1016/j.aca.2013.08.051

To address immunocapture of proteins in large cohorts of clinical samples high throughput sample processing is required. Here a method using the proteomic sample platform, ISET (integrated selective enrichment target) that integrates highly specific immunoaffinity capture of protein biomarker, digestion and sample cleanup with a direct interface to mass spectrometry is presented. The robustness of the on-ISET protein digestion protocol was validated by MALDI MS analysis of model proteins, ranging from 40 fmol to 1 pmol per nanovial. On-ISET digestion and MALDI MS/MS analysis of immunoaffinity captured disease-associated biomarker PSA (prostate specific antigen) from human seminal plasma are presented.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization*
Comparison of MALDI-TOF mass spectrometry with phenotypic methods for identification and characterization of Staphylococcus aureus causing mastitis

Alharbi A, Al-Dubaib M, Elhassan MAS, Elbehiry A

Trop Biomed, 2021 Jun 01;38(2):9-24.
PMID: 33973568 DOI: 10.47665/tb.38.2.032

The emergence of methicillin-resistant Staphylococcus aureus (MRSA) infection is one of the greatest threats to both animal and human health. Our investigation was aimed to identify and differentiate between MRSA and methicillin-sensitive Staphylococcus aureus (MSSA) recovered from mastitic milk using MALDI-TOF mass spectrometry compared with phenotypic methods and studying their susceptibility to various antibiotics. Four hundred milk samples from mastitic animals (cows, sheep, goats, and dromedary camels) were investigated. Phenotypic identification of S. aureus was made through MASTASAPH Latex test, STAPH ID 32, and Vitek 2 system. The proteomic characterization of S. aureus was done by MBT. The Kirby Bauer method was accomplished to detect the resistance of S. aureus strains to antibiotics. The results of the MASTASAPH Latex test, revealed that 54 (46%) were recognized as S. aureus. All S. aureus isolates were identified by MBT with a score of more or equal 2.00. Several peaks were identified in the mass of 4590 Da, 4863 Da, and 4938 Da for MSSA and in the mass of 2636 Da and 3009 Da for MRSA. The MSP dendrogram demonstrated that the S. aureus isolates were classified into one group with a distance level of less or equal 400. The percentage of S. aureus resistance against carbenicillin, erythromycin and kanamycin was 94.4%, 38.88%, and 33.33%, respectively. In conclusion, S. aureus bacteria are among the key triggers for mastitis in Saudi Arabia. MBT is reported to be not only the rapid tool to identify S. aureus but also able to discriminate MRSA from MSSA.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
An unbiased approach for analysis of protein glycosylation and application to influenza vaccine hemagglutinin

An Y, Cipollo JF

Anal Biochem, 2011 Aug 1;415(1):67-80.
PMID: 21545787 DOI: 10.1016/j.ab.2011.04.018

Here a mass spectrometry-based platform for the analysis of glycoproteins is presented. Glycopeptides and released glycans are analyzed, the former by quadrupole orthogonal time-of-flight liquid chromatography/mass spectrometry (QoTOF LC/MS) and the latter by permethylation analysis using matrix-assisted laser desorption/ionization (MALDI)-TOF MS. QoTOF LC/MS analysis reveals the stochastic distribution of glycoforms at occupied sequons, and the latter provides a semiquantitative assessment of overall protein glycosylation. Hydrophilic interaction chromatography (HILIC) was used for unbiased enrichment of glycopeptides and was validated using five model N-glycoproteins bearing a wide array of glycans, including high-mannose, complex, and hybrid subtypes such as sulfo and sialyl forms. Sialyl and especially sulfated glycans are difficult to analyze because these substitutions are labile. The conditions used here allow detection of these compounds quantitatively, intact, and in the context of overall glycosylation. As a test case, we analyzed influenza B/Malaysia/2506/2004 hemagglutinin, a component of the 2006-2007 influenza vaccine. It bears 11 glycosylation sites. Approximately 90% of its glycans are high mannose, and 10% are present as complex and hybrid types, including those with sulfate. The stochastic distribution of glycoforms at glycosylation sites is revealed. This platform should have wide applications to glycoproteins in basic sciences and industry because no apparent bias for any glycoforms is observed.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext Comparative analysis of Mafriwal (Bos taurus × Bos indicus) and Kedah Kelantan (Bos indicus) sperm proteome identifies sperm proteins potentially responsible for higher fertility in a tropical climate

Ashrafzadeh A, Nathan S, Karsani SA

Int J Mol Sci, 2013;14(8):15860-77.
PMID: 23903046 DOI: 10.3390/ijms140815860

The fertility of zebu cattle (Bos indicus) is higher than that of the European purebred (Bos taurus) and crossbred (Bos taurus × Bos indicus) cattle in tropical areas. To identify proteins related to the higher thermo-tolerance and fertility of Zebu cattle, this study was undertaken to identify differences in sperm proteome between the high fertile Malaysian indigenous zebu cattle (Kedah Kelantan) and the sub-fertile crossbred cattle (Mafriwal). Frozen semen from three high performance bulls from each breed were processed to obtain live and pure sperm. Sperm proteins were then extracted, and two-dimensional gel electrophoresis performed to compare proteome profiles. Gel image analysis identified protein spots of interest which were then identified by liquid chromatography mass spectrometry quadrupole time-of-flight (LC MS/MS Q-TOF). STRING network analysis predicted interactions between at least 20 of the identified proteins. Among the identified proteins, a number of motility and energy related proteins were present in greater abundance in Kedah Kelantan. Sperm motility evaluation by Computer Assisted Semen Analysis (CASA) confirmed significantly higher motility in Kedah Kelantan. While results from this study do identify proteins that may be responsible for the higher fertility of Kedah Kelantan, functional characterization of these proteins is warranted to reinforce our understanding of their roles in sperm fertility.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization*
Fulltext Enhanced production, purification, characterization and mechanism of action of salivaricin 9 lantibiotic produced by Streptococcus salivarius NU10

Barbour A, Philip K, Muniandy S

PLoS One, 2013;8(10):e77751.
PMID: 24147072 DOI: 10.1371/journal.pone.0077751

BACKGROUND: Lantibiotics are small lanthionine-containing bacteriocins produced by lactic acid bacteria. Salivaricin 9 is a newly discovered lantibiotic produced by Streptococcus salivarius. In this study we present the mechanism of action of salivaricin 9 and some of its properties. Also we developed new methods to produce and purify the lantibiotic from strain NU10.
METHODOLOGY/PRINCIPAL FINDINGS: Salivaricin 9 was found to be auto-regulated when an induction assay was applied and this finding was used to develop a successful salivaricin 9 production system in liquid medium. A combination of XAD-16 and cation exchange chromatography was used to purify the secondary metabolite which was shown to have a molecular weight of approximately 3000 Da by SDS-PAGE. MALDI-TOF MS analysis indicated the presence of salivaricin 9, a 2560 Da lantibiotic. Salivaricin 9 is a bactericidal molecule targeting the cytoplasmic membrane of sensitive cells. The membrane permeabilization assay showed that salivaricin 9 penetrated the cytoplasmic membrane and induced pore formation which resulted in cell death. The morphological changes of test bacterial strains incubated with salivaricin 9 were visualized using Scanning Electron Microscopy which confirmed a pore forming mechanism of inhibition. Salivaricin 9 retained biological stability when exposed to high temperature (90-100°C) and stayed bioactive at pH ranging 2 to 10. When treated with proteinase K or peptidase, salivaricin 9 lost all antimicrobial activity, while it remained active when treated with lyticase, catalase and certain detergents.
CONCLUSION: The mechanism of antimicrobial action of a newly discovered lantibiotic salivaricin 9 was elucidated in this study. Salivaricin 9 penetrated the cytoplasmic membrane of its targeted cells and induced pore formation. This project has given new insights on lantibiotic peptides produced by S. salivarius isolated from the oral cavities of Malaysian subjects.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Uncovering Tumor-Stroma Inter-relationships Using MALDI Mass Spectrometry Imaging

Boyle ST, Mittal P, Kaur G, Hoffmann P, Samuel MS, Klingler-Hoffmann M

J Proteome Res, 2020 10 02;19(10):4093-4103.
PMID: 32870688 DOI: 10.1021/acs.jproteome.0c00511

Tumorigenesis involves a complex interplay between genetically modified cancer cells and their adjacent normal tissue, the stroma. We used an established breast cancer mouse model to investigate this inter-relationship. Conditional activation of Rho-associated protein kinase (ROCK) in a model of mammary tumorigenesis enhances tumor growth and progression by educating the stroma and enhancing the production and remodeling of the extracellular matrix. We used peptide matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to quantify the proteomic changes occurring within tumors and their stroma in their regular spatial context. Peptides were ranked according to their ability to discriminate between the two groups, using a receiver operating characteristic tool. Peptides were identified by liquid chromatography tandem mass spectrometry, and protein expression was validated by quantitative immunofluorescence using an independent set of tumor samples. We have identified and validated four key proteins upregulated in ROCK-activated mammary tumors relative to those expressing kinase-dead ROCK, namely, collagen I, α-SMA, Rab14, and tubulin-β4. Rab14 and tubulin-β4 are expressed within tumor cells, whereas collagen I is localized within the stroma. α-SMA is predominantly localized within the stroma but is also expressed at higher levels in the epithelia of ROCK-activated tumors. High expression of COL1A, the gene encoding the pro-α 1 chain of collagen, correlates with cancer progression in two human breast cancer genomic data sets, and high expression of COL1A and ACTA2 (the gene encoding α-SMA) are associated with a low survival probability (COLIA, p = 0.00013; ACTA2, p = 0.0076) in estrogen receptor-negative breast cancer patients. To investigate whether ROCK-activated tumor cells cause stromal cancer-associated fibroblasts (CAFs) to upregulate expression of collagen I and α-SMA, we treated CAFs with medium conditioned by primary mammary tumor cells in which ROCK had been activated. This led to abundant production of both proteins in CAFs, clearly highlighting the inter-relationship between tumor cells and CAFs and identifying CAFs as the potential source of high levels of collagen 1 and α-SMA and associated enhancement of tissue stiffness. Our research emphasizes the capacity of MALDI-MSI to quantitatively assess tumor-stroma inter-relationships and to identify potential prognostic factors for cancer progression in human patients, using sophisticated mouse cancer models.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
MALDI Mass Spectrometry Imaging of Early- and Late-Stage Serous Ovarian Cancer Tissue Reveals Stage-Specific N-Glycans

Briggs MT, Condina MR, Ho YY, Everest-Dass AV, Mittal P, Kaur G, et al.

Proteomics, 2019 11;19(21-22):e1800482.
PMID: 31364262 DOI: 10.1002/pmic.201800482

Epithelial ovarian cancer is one of the most fatal gynecological malignancies in adult women. As studies on protein N-glycosylation have extensively reported aberrant patterns in the ovarian cancer tumor microenvironment, obtaining spatial information will uncover tumor-specific N-glycan alterations in ovarian cancer development and progression. matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) is employed to investigate N-glycan distribution on formalin-fixed paraffin-embedded ovarian cancer tissue sections from early- and late-stage patients. Tumor-specific N-glycans are identified and structurally characterized by porous graphitized carbon-liquid chromatography-electrospray ionization-tandem mass spectrometry (PGC-LC-ESI-MS/MS), and then assigned to high-resolution images obtained from MALDI-MSI. Spatial distribution of 14 N-glycans is obtained by MALDI-MSI and 42 N-glycans (including structural and compositional isomers) identified and structurally characterized by LC-MS. The spatial distribution of oligomannose, complex neutral, bisecting, and sialylated N-glycan families are localized to the tumor regions of late-stage ovarian cancer patients relative to early-stage patients. Potential N-glycan diagnostic markers that emerge include the oligomannose structure, (Hex)6 + (Man)3 (GlcNAc)2 , and the complex neutral structure, (Hex)2 (HexNAc)2 (Deoxyhexose)1 + (Man)3 (GlcNAc)2 . The distribution of these markers is evaluated using a tissue microarray of early- and late-stage patients.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Autophagy in Helicobacter pylori Infection and Related Gastric Cancer

Castaño-Rodríguez N, Kaakoush NO, Goh KL, Fock KM, Mitchell HM

Helicobacter, 2015 Oct;20(5):353-69.
PMID: 25664588 DOI: 10.1111/hel.12211

Autophagy, a degradation pathway in which cytoplasmic content is engulfed and degraded by lysosomal hydrolases, plays a pivotal role in infection and inflammation. Given that defects in autophagy lead to increased susceptibility to infection, we investigated the role of autophagy in Helicobacter pylori-related gastric cancer (GC).

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Human neuronal cell protein responses to Nipah virus infection

Chang LY, Ali AR, Hassan SS, AbuBakar S

Virol J, 2007;4:54.
PMID: 17553172

Nipah virus (NiV), a recently discovered zoonotic virus infects and replicates in several human cell types. Its replication in human neuronal cells, however, is less efficient in comparison to other fully susceptible cells. In the present study, the SK-N-MC human neuronal cell protein response to NiV infection is examined using proteomic approaches.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext Characterization of affinity-purified isoforms of Acinetobacter calcoaceticus Y1 glutathione transferases

Chee CS, Tan IK, Alias Z

ScientificWorldJournal, 2014;2014:750317.
PMID: 24892084 DOI: 10.1155/2014/750317

Glutathione transferases (GST) were purified from locally isolated bacteria, Acinetobacter calcoaceticus Y1, by glutathione-affinity chromatography and anion exchange, and their substrate specificities were investigated. SDS-polyacrylamide gel electrophoresis revealed that the purified GST resolved into a single band with a molecular weight (MW) of 23 kDa. 2-dimensional (2-D) gel electrophoresis showed the presence of two isoforms, GST1 (pI 4.5) and GST2 (pI 6.2) with identical MW. GST1 was reactive towards ethacrynic acid, hydrogen peroxide, 1-chloro-2,4-dinitrobenzene, and trans,trans-hepta-2,4-dienal while GST2 was active towards all substrates except hydrogen peroxide. This demonstrated that GST1 possessed peroxidase activity which was absent in GST2. This study also showed that only GST2 was able to conjugate GSH to isoproturon, a herbicide. GST1 and GST2 were suggested to be similar to F0KLY9 (putative glutathione S-transferase) and F0KKB0 (glutathione S-transferase III) of Acinetobacter calcoaceticus strain PHEA-2, respectively.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Identification of a 48kDa tubulin or tubulin-like C6/36 mosquito cells protein that binds dengue virus 2 using mass spectrometry

Chee HY, AbuBakar S

Biochem Biophys Res Commun, 2004 Jul 16;320(1):11-7.
PMID: 15207695

Binding of dengue virus 2 (DENV-2) to C6/36 mosquito cells protein was investigated. A 48 kDa DENV-2-binding C6/36 cells protein (D2BP) was detected in a virus overlay protein-binding assay. The binding occurred only to the C6/36 cells cytosolic protein fraction and it was inhibited by free D2BP. D2BP was shown to bind to DENV-2 E in the far-Western-binding studies and using mass spectrometry (MS) and MS/MS, peptide masses of the D2BP that matched to beta-tubulin and alpha-tubulin chains were identified. These findings suggest that DENV-2 through DENV-2 E binds directly to a 48 kDa tubulin or tubulin-like protein of C6/36 mosquito cells.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods*
Fulltext Francisella philomiragia bacteremia in an immunocompromised patient: a rare case report

Chua HS, Soh YH, Loong SK, AbuBakar S

Ann Clin Microbiol Antimicrob, 2021 Oct 03;20(1):72.
PMID: 34602092 DOI: 10.1186/s12941-021-00475-2

BACKGROUND: Francisella philomiragia is a very rare opportunistic pathogen of humans which causes protean diseases such as pneumonia and other systemic infections. Subsequent failure of prompt treatment may result in poor prognosis with mortality among infected patients.
CASE PRESENTATION: The present report describes a case of F. philomiragia bacteraemia first reported in Malaysia and Asian in a 60-year-old patient with underlying end-stage renal disease (ESRF) and diabetes mellitus. He presented with Acute Pulmonary Oedema with Non-ST-Elevation Myocardial Infarction (NSTEMI) in our hospital. He was intubated in view of persistent type I respiratory failure and persistent desaturation despite post haemodialysis. Blood investigation indicated the presence of ongoing infection and inflammation. The aerobic blood culture growth of F. philomiragia was identified using the matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (Score value: 2.16) and confirmed by 16S Ribosomal DNA (16S rDNA) sequencing. He was discharged well on day 26 of admission, after completing one week of piperacillin/tazobactam and two weeks of doxycycline.
CONCLUSION: Clinical suspicion should be raised if patients with known risk factors are presenting with pneumonia or pulmonary nodules especially as these are the most common manifestations of F. philomiragia infection. Early diagnosis via accurate laboratory identification of the organism through MALDI-TOF mass spectrometry and molecular technique such as 16S rDNA sequencing are vital for prompt treatment that results in better outcomes for the afflicted patients.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Proteomic analysis reveals that treatment with tocotrienols reverses the effect of H₂O₂ exposure on peroxiredoxin expression in human lymphocytes from young and old individuals

Dahlan HM, Karsani SA, Rahman MA, Hamid NA, Top AG, Ngah WZ

J Nutr Biochem, 2012 Jul;23(7):741-51.
PMID: 21840697 DOI: 10.1016/j.jnutbio.2011.03.018

Vitamin E has been suggested to modulate age-associated changes by altering the redox balance resulting in altered gene and/or protein expression. Here we have utilized proteomics to determine whether such regulation in protein expression occurs in human lymphocytes from two different age groups stressed with H₂O₂ and then treated with vitamin E in the form of tocotrienol-rich fraction (TRF). In this study, lymphocytes obtained from young (30-49 years old) and old (>50 years old) volunteers were first challenged with 1 mM H₂O₂. They were then treated by exposure to 50, 100 and 200 μg/ml TRF. Two-dimensional gel electrophoresis followed by MALDI-TOF/TOF (matrix-assisted laser desorption/ionization time-of-flight/time-of-flight) tandem mass spectrometry was then performed on whole-cell protein extracts to identify proteins that have changed in expression. A total of 24 proteins were found to be affected by H₂O₂ and/or TRF treatment. These included proteins that were related to metabolism, antioxidants, structural proteins, protein degradation and signal transduction. Of particular interest was the regulation of a number of proteins involved in stress response--peroxiredoxin-2, peroxiredoxin-3 and peroxiredoxin-6-all of which were shown to be down-regulated with H₂O₂ exposure. The effect was reversed following TRF treatment. The expression of peroxiredoxin-2 and peroxiredoxin-6 was confirmed by quantitative reverse transcriptase polymerase chain reaction. These results suggested that TRF directly influenced the expression dynamics of the peroxiredoxin-2, thus improving the cells ability to resist damage caused by oxidative stress.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
Patients with nasopharyngeal carcinoma demonstrate enhanced serum and tissue ceruloplasmin expression

Doustjalali SR, Yusof R, Govindasamy GK, Bustam AZ, Pillay B, Hashim OH

J. Med. Invest., 2006 Feb;53(1-2):20-8.
PMID: 16537992

The proteomics approach was adopted to study the simultaneous expression of serum proteins in patients with nasopharyngeal carcinoma (NPC). We have subjected unfractionated whole sera of ten newly diagnosed Malaysian Chinese patients with WHO type III NPC to two-dimensional gel electrophoresis (2-DE) and image analysis. The results obtained were then compared to that generated from sera of ten normal healthy controls of the same ethnic group and range of age. Our data demonstrated that the serum high abundance 2-DE protein profiles of NPC patients were generally similar to that of the controls, with exception of the ceruloplasmin (CPL) spots (identified by mass spectrometric analysis and MASCOT database search), which showed higher expression. The enhanced expression of CPL in the patients' sera was confirmed by competitive ELISA. Immunohistochemical analysis of nasopharyngeal lesions of NPC patients demonstrated moderate to strong positive CPL staining in the cytoplasm of cells at the regions of malignancy but only weak cytoplasmic staining at normal epithelial lining areas. When follow-up 2-DE and ELISA studies were performed on five of the NPC patients who responded positively to six months treatment, the difference in CPL expression was no longer significant.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext Quorum sensing activity in Pandoraea pnomenusa RB38

Ee R, Lim YL, Kin LX, Yin WF, Chan KG

Sensors (Basel), 2014;14(6):10177-86.
PMID: 24919016 DOI: 10.3390/s140610177

Strain RB38 was recovered from a former dumping area in Malaysia. MALDI-TOF mass spectrometry and genomic analysis identified strain RB-38 as Pandoraea pnomenusa. Various biosensors confirmed its quorum sensing properties. High resolution triple quadrupole liquid chromatography-mass spectrometry analysis was subsequently used to characterize the N-acyl homoserine lactone production profile of P. pnomenusa strain RB38, which validated that this isolate produced N-octanoyl homoserine lactone as a quorum sensing molecule. This is the first report of the production of N-octanoyl homoserine lactone by P. pnomenusa strain RB38.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext Quorum sensing activity of Serratia fonticola strain RB-25 isolated from an ex-landfill site

Ee R, Lim YL, Tee KK, Yin WF, Chan KG

Sensors (Basel), 2014 Mar 12;14(3):5136-46.
PMID: 24625739 DOI: 10.3390/s140305136

Quorum sensing is a unique bacterial communication system which permits bacteria to synchronize their behaviour in accordance with the population density. The operation of this communication network involves the use of diffusible autoinducer molecules, termed N-acylhomoserine lactones (AHLs). Serratia spp. are well known for their use of quorum sensing to regulate the expression of various genes. In this study, we aimed to characterized the AHL production of a bacterium designated as strain RB-25 isolated from a former domestic waste landfill site. It was identified as Serratia fonticola using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry analysis and this was confirmed by 16S ribosomal DNA sequencing. High resolution triple quadrupole liquid chromatography-mass spectrometry analysis of S. fonticola strain RB-25 spent culture supernatant indicated the existence of three AHLs namely: N-butyryl-L-homoserine lactone (C4-HSL), N-hexanoyl-L-homoserine lactone (C6-HSL) and N-(3-oxohexanoyl) homoserine-lactone (3-oxo-C6 HSL). This is the first report of the production of these AHLs in S. fonticola.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Fulltext Proteomic analysis of saliva identifies potential biomarkers for orthodontic tooth movement

Ellias MF, Zainal Ariffin SH, Karsani SA, Abdul Rahman M, Senafi S, Megat Abdul Wahab R

ScientificWorldJournal, 2012;2012:647240.
PMID: 22919344 DOI: 10.1100/2012/647240

Orthodontic treatment has been shown to induce inflammation, followed by bone remodelling in the periodontium. These processes trigger the secretion of various proteins and enzymes into the saliva. This study aims to identify salivary proteins that change in expression during orthodontic tooth movement. These differentially expressed proteins can potentially serve as protein biomarkers for the monitoring of orthodontic treatment and tooth movement. Whole saliva from three healthy female subjects were collected before force application using fixed appliance and at 14 days after 0.014'' Niti wire was applied. Salivary proteins were resolved using two-dimensional gel electrophoresis (2DE) over a pH range of 3-10, and the resulting proteome profiles were compared. Differentially expressed protein spots were then identified by MALDI-TOF/TOF tandem mass spectrometry. Nine proteins were found to be differentially expressed; however, only eight were identified by MALDI-TOF/TOF. Four of these proteins-Protein S100-A9, immunoglobulin J chain, Ig alpha-1 chain C region, and CRISP-3-have known roles in inflammation and bone resorption.

Matched MeSH terms: Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

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