Lysosomes are composed of soluble and transmembrane proteins that target lysosomes in a signal-dependent manner. Our engineers use a wide range of methods including subcellular isolation, liquid chromatography tandem mass spectrometry (LC-MS/MS) for lysosomal protein identification. Here, CD BioSciences provides specialized lysosomal membrane proteins identification services to support lysosomal proteomic studies.
Background of Lysosomal Membrane Proteins Identification
Lysosomal membranes primarily act as a physical barrier separating the tubular acidic environment from the cytoplasmic environment and also play an important role in many cellular events. To date, more than 120 lysosomal membrane proteins (LMPs) have been associated with lysosomal functions, including acidification, transport of metabolites and ions across the membrane, intracellular transport of hydrolases, and regulation of membrane fusion events. LMPs are not modified by mannose-6-phosphate (Man6-P) and therefore are not dependent on Man6-P receptors for classification. LMPs are usually highly glycosylated and may form continuous glycoproteins on the luminal side of the lysosomal membrane. The most abundant type 1 transmembrane proteins in the lysosomal membrane are the lysosomal-associated membrane proteins LAMP-1 and LAMP-2, using more than 10 glycosylation sites. These proteins serve as the core of lysosomal biogenesis and they become attractive targets for studying the mechanisms of lysosomal regulation in lysosomal diseases.
Fig. 1. Major functions of lysosomal membrane proteins. (Saftig P, et al., 2009)
Our Lysosomal Membrane Proteins Identification Services
Soluble lysosomal proteins have been extensively studied, but knowledge about lysosomal membrane proteins is still quite limited, despite the multiple and critical functions of membranes. Currently, CD BioSciences is focused on studying the composition of lysosomal proteins using large-scale mass spectrometry (MS)-based methods. Here, our engineers provide satisfactory lysosomal membrane protein identification services by combining subcellular hierarchical separation, differential extraction and protein isolation techniques with proteomic protein identification.
Our goal is to determine the complex composition and function of proteins in the intact membrane fraction of lysosomes. We have successfully established a master list of thousands of gene products from both lysosomal-rich and non-lysosomal-rich fractions. To extend our lysosomal protein identification capabilities, we have focused specifically on new potential lysosomal transporter proteins and adopted the following strategy.
- MS analysis prior to combining subcellular and biochemical isolation techniques to enrich for lysosomes.
- Analysis of lysosomal membrane fractions using LC-MS/MS.
- Developing a comparative proteomic analysis tool based on spectral counting.
- Lysosomal localization by overexpression of epitope-tagged fusion proteins.
Our Advantages
- Advanced technical support, such as 2D-PAGE, strong cation exchange chromatography (SCX), SDS-PAGE.
- Allows identification of novel resident lysosomal membrane proteins, such as transporter proteins.
- Coupled display of selective lysosomal density changes and MS quantification allows simultaneous identification and validation of lysosomal candidates.
- Contributes to a deeper understanding of the characterization of lysosomal membranes and their associated proteins.
- Mature platform for comparative, semi-quantitative proteomics analysis.
- Allows the use of spectral counting to assess the relative abundance of proteins.
We look forward to collaborating with you. You are always welcome to engage in discussions with us at any point of the project. If you are interested in our services, please feel free to contact us for more information.
Reference
- Saftig P, Klumperman J. (2009) Lysosome biogenesis and lysosomal membrane proteins: trafficking meets function[J]. Nature reviews Molecular cell biology. 10(9): 623-635.