Hexokinase is the first enzyme in the glycolytic pathway, catalyzing the transfer of a phosphoryl group from ATP to glucose to form glucose-6-phosphate and ADP. In mammals, four distinct enzymes-types 1 to 4 hexokinases-have been identified. The enzyme is found in most cells, but there is tissue specificity for the particular type of hexokinase. Hexokinase2 is found in the skeletal muscle and includes hydrophobic N-terminal sequence capable of targeting the hexokinase to mitochondria. Recombinant human Hexokinase2, fused to His tag at N-terminus, was expressed in E. coli and purified by using conventional chromatography techniques. Specific Activity: 3-4 units/ml obtained by measuring the increase of NADPH in absorbance at 340nm resulting from the reduction of NADP. In the coupled mode, one unit will produce 1.0umole of NADPH per minute as glucose is phosphorylated by ATP at pH 7.4 at 25C. Activity Assay: 1. Prepare a 2.57ml reaction mixture into a suitable container: The final concentrations are 39mM triethanolamine, 216 mM D-glucose, 0.74mM ATP, 7.8mM MgCl2, 1.1mM beta-NADP, 2.5units G6PD. 2. Equilibrate to 25C and monitor at A340nm until the value is constant using a spectrophotometer. 3. Add 5ug of H2040-52 into reaction mixture and mix immediately. 4. Record the increase at A340nm for 5 minutes. Amino Acid Sequences: MGSSHHHHHH/ SSGLVPRGSH/ MIASHLLAYF/ FTELNHDQVQ/ KVDQYLYHMR/ LSDETLLEIS/ KRFRKEMEKG/ LGATTHPTAA/ VKMLPTFVRS/ TPDGTEHGEF/ LALDLGGTNF/ RVLWVKVTDN/ GLQKVEMENQ/ IYAIPEDIMR/ GSGTQLFDHI/ AECLANFMDK/ LQIKDKKLPL/ GFTFSFPCHQ/ TKLDESFLVS/ WTKGFKSSGV/ EGRDVVALIR/ KAIQRRGDFD/ IDIVAVVNDT/ VGTMMTCGYD/ DHNCEIGLIV/ GTGSNACYME/ EMRHIDMVEG/ DEGRMCINME/ WGAFGDDGSL/ NDIRTEFDQE/ IDMGSLNPGK/ QLFEKMISGM/ YMGELVRLIL/ VKMAKEELLF/ GGKLSPELLN/ TGRFETKDIS/ DIEGEKDGIR/ KAREVLMRLG/ LDPTQEDCVA/ THRICQIVST/ RSASLCAATL/ AAVLQRIKEN/ KGEERLRSTI/ GVDGSVYKKH/ PHFAKRLHKT/ VRRLVPGCDV/ RFLRSEDGSG/ KGAAMVTAVA/ YRLADQHRAR/ QKTLEHLQLS/ HDQLLEVKRR/ MKVEMERGLS/ KETHASAPVK/ MLPTYVCATP/ DGTEKGDFLA/ LDLGGTNFRV/ LLVRVRNGKW/ GGVEMHNKIY/ AIPQEVMHGT/ GDELFDHIVQ/ CIADFLEYMG/ MKGVSLPLGF/ TFSFPCQQNS/ LDESILLKWT/ KGFKASGCEG/ EDVVTLLKEA/ IHRREEFDLD/ VVAVVNDTVG/ TMMTCGFEDP/ HCEVGLIVGT/ GSNACYMEEM/ RNVELVEGEE/ GRMCVNMEWG/ AFGDNGCLDD/ FRTEFDVAVD/ ELSLNPGKQR/ FEKMISGMYL/ GEIVRNILID/ FTKRGLLFRG/ RISERLKTRG/ IFETKFLSQI/ ESDCLALLQV/ RAILQHLGLE/ STCDDSIIVK/ EVCTVVARRA/ AQLCGAGMAA/ VVDRIRENRG/ LDALKVTVGV/ DGTLYKLHPH/ FAKVMHETVK/ DLAPKCDVSF/ LQSEDGSGKG/ AALITAVACR/ IREAGQR Storage and Stability: May be stored at 4C for short-term only. Aliquot to avoid repeated freezing and thawing.. Store at -20C. Aliquots are stable for 6 months after receipt at -20C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
Molekulargewicht:
104.1
Reinheit:
90% by SDS PAGE
Formulierung:
Supplied as a liquid in 20mM Tris-HCl, pH8.0, 10% glycerol
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