Expression in Escherichia coli and purification of hexahistidine-tagged human tissue transglutaminase.
Recent evidence suggests that aberrant transglutaminase activity is associated with a wide variety of diseases. Tissue transglutaminase is the most widely distributed of the six well-characterized transglutaminases in humans. We describe a method for expressing hexahistidine-tagged human tissue transglutaminase in Escherichia coli BL21(DE3) using the pET-30 Ek/LIC expression vector. Purification of the expressed enzyme from suspensions of E. coli cells treated with CelLytic B Bacterial Cell Lysis/Extraction Reagent was accomplished by immobilized metal (Ni2+) affinity column chromatography. The procedure typically yields highly purified and highly active recombinant human tissue transglutaminase in about 1 day (about 0.6 mg/from a 1-liter culture).
Shi
Qingli
Q
Department of Neurology and Neuroscience, Weill Medical College of Cornell University, New York, NY 10021, USA.
Kim
Soo-Youl
SY
Blass
John P
JP
Cooper
Arthur J L
AJ
D004795:Enzyme Stability; D004926:Escherichia coli; D006639:Histidine / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011503:Transglutaminases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*
D011994:Recombinant Proteins; D006639:Histidine; D011503:Transglutaminases