Glycosylation is the most common and important form of post-translational modification. pastoris can N-glycosylate proteins via mannose oligosaccharide linked to asparagine through two N-acetylglucosamines. Pichia pastoris is a widely used industrial methylotrophic yeast that has been developed as a useful experimental tool in protein engineering and production. Our report may also provide theoretical support for the improvement of enzyme expression and stability based on the N-linked glycosylation modification to meet the future needs of the biotechnological industry. The N-glycans of RCL on the different sites had different functions for the secretion and enzymatic properties of the lipase. RCL was N-glycosylated when expressed in P. Moreover, the thermostability analysis of RCL revealed that the lipase with more N-glycan was more thermostable. On the other hand, the little amount of N-glycan on N-48 had no effect both on the secretion and activity of RCL in P. Although the N-glycan on N-14 had no effect on the secretion of RCL, this glycan was beneficial for the lipase catalytic activity. RT-PCR results showed that the mRNA level of proRCLCN60Q remained unchanged even though the protein secretion was hampered. The glycan on N-60 played a key role in the expression and secretion of RCL. The majority of the sites N-14 and N-60 were glycosylated, but the glycosylation degree of the site N-48 was only a very small portion. In this study, we demonstrated that RCL expressed in Pichia pastoris was N-glycosylated at the sites N-14, N-48 and N-60. The aim of the present study was to determine whether RCL undergoes asparagine-linked (N-linked) glycosylation and to examine the role of this modification in RCL expression and function. Rhizopus chinensis lipase (RCL) is one of the most important industrial lipases, and it has four potential N-linked glycosylation sites. pastoris have attracted increasing attention from scholars. In recent years, glycosylation studies in P. It is common for proteins expressed in P. For example, the enhanced standard may be necessary when using a 1:50,000 (25 ng/mL) to 1:100,000 (10 ng/mL) dilution of secondary antibody.The methylotrophic yeast, Pichia pastoris, is widely used as a useful experimental tool in protein engineering and production. SuperSignal Enhanced Molecular Weight Protein ladder is formulated specifically for applications requiring mouse monoclonal antibodies and experiments where very dilute antibody concentrations are used. This formulation is appropriate for use with most rabbit and other non-mouse polyclonal antibodies. Two versions of the MW marker are offered. The protein markers can then be visualized using appropriate substrates for enzyme-labeled antibodies or via fluorescent dye-labeled antibodies (not recommended for antibodies labeled with fluors in the 500–550 nm channel). The ladder’s recombinant proteins bind to the antibodies used in the western blot through the IgG binding site. The protein standard is supplied in a ready-to-use format for direct loading onto gels no need to heat, reduce, or add sample buffer prior to use.Ĭompare and view all other protein standards and ladders › Thermo Scientific SuperSignal Molecular Weight Protein Ladder (20 to 150K) is a mix of eight unstained recombinant proteins with IgG-binding sites for chemiluminescent, fluorescent, chromogenic or other detection systems.
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