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Liverpool Bio-Innovation Hub 

William Henry Duncan Building

6 West Derby Street

Liverpool

L7 8TX

United Kingdom

 

t: +44 (0) 151 794 9317 

e: orders@geminibioscience.com 

    info@geminibioscience.com 

Copyright © 2015 Gemini Biosciences Ltd

PRODUCTS AND SERVICES

1 PROTEOMICS
1.4 Post translational modifications
I. PHOSPHOPROTEOMICS AND PHOSPHORYLATION SITE MAPPING

 

(i) Quantitative phosphoproteomics

We offer a comprehensive phosphoproteomics service to our clients that includes experimental design, advice with sample preparation, fractionation, affinity enrichment of phosphopeptides using TiO2, phospho Serine/Threonine or phospho Tyrosine antibodies, LC-MS/MS, data analysis and interpretation. A final project report can be prepared at completion of each project according to the customer's needs.

Our quantitative phosphoproteomics analysis could either be done using metabolically labeled samples (using SILAC media), chemically labeled samples (iTRAQ) or a label-free quantitative proteomics approach depending on the nature of the customers samples.

 

Please contact us as early as possible during the planning stages of your project so that we can discuss the best approach that meets the requirements of your project.

 

(ii) Phosphorylation site mapping

Our state of the art mass spectrometry equipment can be used to identify sites of phosphorylation from a protein in a 1D band or 2D gel spot. The 1D band or 2D spot must have good intensity by colloidal Coomassie staining in order to proceed; this gives us a better chance of identifying the sites of phosphorylation as the level of protein usually gives no indication of the stoichiometry of the phosphorylation present. Samples are normally processed and then in-gel digested with a protease e.g. trypsin, chymotrysin, etc. The digested samples are then extracted and are taken through three stages of analysis.

 

The stages include:

a) Identification of protein(s) by high-resolution mass spectrometry

b) Detection of phosphorylated peptides

c) Data interpretation and final reporting

Mascot reports of protein identification(s) and identification of sites of phosphorylation are provided electronically or as a hardcopy if requested.

 

 

II. METHYLATION PROTEOMICS

Gemini Biosciences methylation proteomics service allows the customer to obtain information on the methylation status of their biological samples. Our workflow involves using mono-methylated arginine specific antibodies to enrich for modified peptides in protease digested cell or tissue extracts. The antibody-purified modified peptides are then analysed by LC-MS/MS to determine modified sites and quantification of peptides/proteins to obtain quantitative profiles of hundreds of modified proteins in the customer's samples.

 

 

III. ACETYLATION PROTEOMICS

Our acetylation proteomics service provides the customer with information on the acethylation status of their biological samples. Our workflow involves using high affinity acetylated-lysine (Ac-K) specific antibodies to enrich for acetyl-modified peptides in protease digested cell or tissue extracts. The antibody-purified modified peptides are then analysed by LC-MS/MS to determine modified sites and quantification of peptides/proteins to obtain quantitative profiles of hundreds of modified proteins in the customer's samples.

 

 

IV. UBIQUITINATION PROTEOMICS

Ubiquitin modification of cellular proteins is one of the major regulatory steps that directs protein degradation through regulation of proteasome activity. Since in most instances ubiquitination are the primary event in target selection, the system of ubiquitination and deubiquitination might be of similar importance. Furthermore, protein ubiquitination is involved in functions different from targeting proteins for degradation. Therefore, depending on the nature of the modification i.e. mono or polyubiquitination and on how ubiquitin chains are linked together, post-translational modifications of cellular proteins by covalent attachment of ubiquitin and ubiquitin-like proteins are involved in transcriptional regulation, receptor internalization, DNA repair, stabilization of protein complexes and autophagy. 

 

The ubiquitin proteomics service offered by Gemini Bio uses the K-GG motif specific antibody to enrich for ubiquitinated substrates by pulling down ubiquitinated peptides in protease digested cell or tissue extracts. The antibody-purified modified peptides are then analysed by LC-MS/MS to determine modified sites and quantification of peptides/proteins to obtain quantitative profiles of the modified proteins in the customer's samples.