Proteomic Profiling and Protein Identification by MALDI-TOF Mass Spectrometry in Unsequenced Parasitic Nematodes
News Apr 16, 2012
Lack of genomic sequence data and the relatively high cost of tandem mass spectrometry have hampered proteomic investigations into helminths, such as resolving the mechanism underpinning globally reported anthelmintic resistance. Whilst detailed mechanisms of resistance remain unknown for the majority of drug-parasite interactions, gene mutations and changes in gene and protein expression are proposed key aspects of resistance. Comparative proteomic analysis of drug-resistant and -susceptible nematodes may reveal protein profiles reflecting drug-related phenotypes. Using the gastro-intestinal nematode, Haemonchus contortus as case study, we report the application of freely available expressed sequence tag (EST) datasets to support proteomic studies in unsequenced nematodes. EST datasets were translated to theoretical protein sequences to generate a searchable database. In conjunction with matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF-MS), Peptide Mass Fingerprint (PMF) searching of databases enabled a cost-effective protein identification strategy. The effectiveness of this approach was verified in comparison with MS/MS de novo sequencing with searching of the same EST protein database and subsequent searches of the NCBInr protein database using the Basic Local Alignment Search Tool (BLAST) to provide protein annotation. Of 100 proteins from 2-DE gel spots, 62 were identified by MALDI-TOF-MS and PMF searching of the EST database. Twenty randomly selected spots were analysed by electrospray MS/MS and MASCOT Ion Searches of the same database. The resulting sequences were subjected to BLAST searches of the NCBI protein database to provide annotation of the proteins and confirm concordance in protein identity from both approaches. Further confirmation of protein identifications from the MS/MS data were obtained by de novo sequencing of peptides, followed by FASTS algorithm searches of the EST putative protein database. This study demonstrates the cost-effective use of available EST databases and inexpensive, accessible MALDI-TOF MS in conjunction with PMF for reliable protein identification in unsequenced organisms.
The article is published online in the journal PLoS ONE and is free to access.
'Royal' Pheromone Identified in TermitesNews
Researchers have for the first time identified a specific chemical used by the higher termite castes – the queens and the kings – to communicate their royal status with worker termites. The findings could advance knowledge of termite evolution, behavior and control.READ MORE
Making Fragrances Last LongerNews
From floral perfume to fruity body wash and shampoos, scents heavily influence consumer purchases. But for most, the smell doesn’t last long after showering before it fades away. Scientists have now developed a way to get those fragrances to stick to the skin longer instead of washing down the drain immediately after being applied.READ MORE
Auto Industry Tip Improves Smoked FoodNews
Infusing foods with smoke can impart delicious flavors, but could also come with an unwelcome side of carcinogens. To reduce the carcinogen content of smoked foods, researchers took a lesson from the automobile industry, running the smoke through a zeolite filter to remove harmful compounds. It worked, and with a happy bonus: superior smoke flavor.READ MORE
Comments | 0 ADD COMMENT
11th International Conference and Exhibition on Metabolomics & Systems Biology
May 17 - May 19, 2018
6th Annual Congress on Biology and Medicine of Molecules
Sep 17 - Sep 18, 2018