How to purify SUMO Protease?
How to purify SUMO Protease?
SUMO Protease is purified from E. coli by affinity chromatography using the polyhistidine tag. Store SUMO Protease at -20°C (after first-time use) or at -80°C for long-term storage. Avoid multiple freeze/thaw cycles at -80°C.
What is SUMO Protease?
SUMO Protease, also known as Ulp, is a recombinant fragment of ULP1 (Ubl-specific protease 1) from Saccharomyces cerevisiae. It is highly specific for the SUMO protein fusion, recognizing the tertiary structure of SUMO rather than an amino acid sequence.
How do I get rid of sumo tag?
SUMO fusion proteins can be cleaved to remove the SUMO moiety using SUMO-specific proteases such as Ulp1. Here, we describe the use of vectors for the expression of recombinant proteins in E. coli as fusions to the Drosophila SUMO protein.
How do you use SUMO protease?
Add 1 unit of protease per 10-100 µg of fusion protein. Incubate at 30 °C for 1 hour, or at 4 °C overnight. To evaluate cleavage, remove a sample of the reaction (5-10 µg of fusion protein).
How do I get rid of SUMO tag?
What is the purpose of a SUMO tag?
SUMO Tag Definition Sumo tag is most frequently used as N-end fusion sequence in yeast to increase the expression and solubility of the desired recombinant protein. SUMO proteins are similar to ubiquitin in their folded structure but possess only about 20% homology to the amino acid sequence of ubiquitin.
What is protease activity?
A protease (also called a peptidase or proteinase) is an enzyme that catalyzes (increases reaction rate or “speeds up”) proteolysis, the breakdown of proteins into smaller polypeptides or single amino acids. They do this by cleaving the peptide bonds within proteins by hydrolysis, a reaction where water breaks bonds.
How does a SUMO tag work?
SUMO enhances recombinant protein expression and solubility in yeast. DeSUMOylases cysteine proteases used to remove SUMO (for Small Ubiquitin-like MOdifier) tags, are efficient and precise. They enable the release of recombinant proteins with the desired N-terminal amino acid.
What are examples of proteases?
The three main proteolytic enzymes produced naturally in your digestive system are pepsin, trypsin and chymotrypsin. Your body produces them to help break down dietary proteins like meat, eggs and fish into smaller fragments called amino acids. These can then be properly absorbed and digested.
Why do we need protease?
Proteases play many vital roles in cellular processes, including blood coagulation, food digestion, apoptosis, and autophagy. These processes are essential and, thus, proteases must work efficiently to ensure the survival of organisms.
How big is the SUMO tag?
Structure. SUMO proteins are small; most are around 100 amino acids in length and 12 kDa in mass. The exact length and mass varies between SUMO family members and depends on which organism the protein comes from.
Are proteases good or bad?
Proteolytic enzymes are generally considered safe but can cause side effects in some people. It’s possible you may experience digestive issues like diarrhea, nausea and vomiting, especially if you take very high doses (34).