Granzyme B is a cysteine protease found in the cytoplasmic granules of cytolytic T lymphocytes (CTL) and natural killer (NK) cells. Granzyme B is required for the induction of target cell lysis, which occurs as part of cell-mediated immune responses, and can activate apoptosis in target cells by both caspase-dependent and caspase-independent mechanisms. Proteolytic cleavage of substrates by Granzyme B takes place primarily after aspartic acid residues. Recombinant Murine Granzyme B is a glycosylated 227 amino acid protein, comprising the mature active portion of the murine Granzyme B precursor. The apparent molecular weight is 28.9 kDa by mass spectrometry.
Amino Acid Sequence
Determined by its ability to cleave a synthetic chromogentic Granzyme B substrate. The expected specific activity, when using the Ac-IEPD-pNA substrate at 25οC, is greater than 750 nM/min per μg of enzyme.
Endotoxin level is <0.1 ng/μg of protein (<1EU/μg).
Centrifuge the vial prior to opening. Reconstitute in water to a concentration of 0.1–1.0 mg/ml. Do not vortex. For extended storage, it is recommended to further dilute in a buffer containing a carrier protein (example 0.1% BSA) and store in working aliquots at –20°C to –80°C
Apoptosis, Immune Response, Infection and Inflammation, Transplantation
Granzymes are exogenous serine proteinases (enzymes) that are released from cytoplasmic granules of cytotoxic lymphocytes (CTLs) and NK cells. The name “granzymes” is derived from: granules + enzymes. These granules contain next to granzymes other proteins including a pore-forming protein (Perforin). Upon binding of the CTL to a target cell (by CTL-receptor and antigen-presenting MHC molecules on the target cell) the contents of the granules are released in the intercellular space where after perforine will “perforate” the target cell membrane by forming transmembrane pores. Through these pores the granzymes can now enter the cytosol of the target cell. Granzyme B activates the intracellular cascade of caspases finally resulting in the killing of the target cells. Also Granzyme A is able to induce apoptosis in the target cell but the molecular mechanisms of the pathway involved need to be clarified. Percentages of the Granzyme A and B positive CTLs can be determined by flow cytometry and immunocytochemical methods for many disorders. Not all granzymes enter the target cell, part of them also “leak” in to the peripheral blood and other biological fluids. Detectable amounts of granzymes have been found to circulate in healthy volunteers. These soluble granzymes can be measured by ELISAs. Viral infections: Increased levels of soluble granzymes have been found with patients suspected of an increased NK cell and CTL-response caused by systemic viral infections such as EBV, HIV, CMV, hepatitis A and Dengue fever. Lymphomas and carcinomas: It is shown that the presence of a high percentage of Granzyme B positive CTLs in glands of patients suffering from Hodgkin’s disease correlate with a severe prognosis. Rheumatoid arthritis: Soluble Granzyme A and B is increased in synovial fluid from rheumatoid arthritis and significantly higher than levels in patients with osteoarthrosis. Transplantation: Granzymes are likely involved in the acute rejection of kidney-transplants, as infiltrating lymphocytes in the rejected kidney strongly express granzymes. Increasing plasma levels of soluble granzymes in patients with a kidney transplants suggest a systemic viral infection, in particular an infection by CMV.