Expression of recombinant rat myo-inositol 1,4,5-trisphosphate 3-kinase B suggests a regulatory role for its N-terminus.
We have expressed rat myo-inositol 1,4,5-trisphosphate (IP3) 3-kinase B as both a full-length, recombinant, non-fusion protein and a full-length, recombinant, fusion protein with maltose-binding protein (MBP) in Escherichia coli. The fusion protein with MBP is soluble, binds calmodulin and is enzymically active whereas the non-fusion protein is insoluble and does not bind calmodulin unless co-expressed with bacterial chaperone proteins (either GroES and GroEL, or DnaK, DnaJ and GrpE). However, soluble, calmodulin-binding non-fusion IP3 3-kinase B is enzymically inactive. The catalytic domain of the enzyme has previously been shown to reside near the C-terminus; the results we present suggest an auto-regulatory role for the N-terminus.
Thomas
S
S
Department of Biochemistry, University of Bristol, U.K.
Soriano
S
S
d'Santos
C
C
Banting
G
G
D018528:ATP-Binding Cassette Transporters; D000818:Animals; D001665:Binding Sites; D002147:Calmodulin / Q000378:metabolism; D002352:Carrier Proteins / Q000096:biosynthesis; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli; D029968:Escherichia coli Proteins; D007700:Kinetics; D058619:Maltose-Binding Proteins; D018832:Molecular Chaperones / Q000096:biosynthesis / Q000378:metabolism; D009004:Monosaccharide Transport Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; D002147:Calmodulin; D002352:Carrier Proteins; D017931:DNA Primers; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D018832:Molecular Chaperones; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins; C461599:maltose transport system, E coli; D017853:Phosphotransferases (Alcohol Group Acceptor); C051038:Inositol 1,4,5-trisphosphate 3-kinase