Purification and Quality Control
The His-tag recombinant protein is purified by affinity chromatography in combination with FPLC columns. The purified SNARK (K85R mutant) is greater than 90% homogeneous based on SDS-PAGE analysis.

Unit Definition (Activity)
1 unit equals 1 nanogram of purified protein.

Applications
SNARK is capable of autophosphorylation in vitro; immunoprecipitated SNARK has been known to show phosphotransferase activity with a synthetic peptide substrate. 10ng of SNARK is sufficient to phosphorylate 1ug of heat-inactivated nuclear or cytoplasmic extract at 30oC for 30 min.

Formulation and Storage
The protein is in 20mM Tris-HCl pH7.9,100mM NaCl, 0.2mM EDTA, 1mM DTT and 20% glycerol. Stored at -70°C before use. Avoid repeated freeze thaw cycles.

Synonym
Homo sapiens NUAK family, SNF1-like kinase, 2 (NUAK2); NUAK2; DKFZp434J037; DKFZp686F01113; FLJ90349 and SNARK.

Protein Sequence
MESLVFARRS GPTPSAAELA RPLAEGLIKS PKPLMKKQAV KRHHHKHNLR HRYEFLETLG
KGTYGKVKKA RESSGRLVAI KSIRRDKIKD EQDLMHIRRE IEIMSSLNHP HIIAIHEVFE
NSSKIVIVME YASRGDLYDY ISERQQLSER EARHFFRQIV SAVHYCHQNR VVHRDLKLEN
ILLDANGNIK IADFGLSNLY HQGKFLQTFC GSPLYASPEI VNGKPYTGPE VDSWSLGVLL
YILVHGTMPF DGHDHKILVK QISNGAYREP PKPSDACGLI RWLLMVNPTR RATLEDVASH
WWVNWGYATR VGEQEAPHEG GHPGSDSARA SMADWLRRSS RPLLENGAKV CSFFKQHAPG
GGSTTPGLER QHSLKKSRKE NDMAQSLHSD TADDTAHRPG KSNLKLPKGI LKKKVSASAE
GVQEDPPELS PIPASPGQAA PLLPKKGILK KPRQRESGYY SSPEPSESGE LLDAGDVFVS
GDPKEQKPPQ ASGLLLHRKG ILKLNGKFSQ TALELAAPTT FGSLDELAPP RPLARASRPS
GAVSEDSILS SESFDQLDLP ERLPEPPLRG CVSVDNLTGL EEPPSEGPGS CLRRWRQDPL
GDSCFSLTDC QEVTATYRQA LRVCSKLT

Background
Highest expression of SNARK is found in kidney, followed in order by brain, ovary, heart, lung, skeletal muscle, pancreas, testis, and spleen. (1) In insulin-stimulated adipocytes, SIK2 phosphorylates IRS1 at ser794 and may modulate the efficiency of insulin signal transduction, eventually causing the insulin resistance associated with diabetes. (2) Cells with reduced expression of SNARK also showed reduced motility and invasiveness in response to CD95 engagement. SNARK therefore represents an NF-kappaB-regulated anti-apoptotic gene that contributes to the tumor-promoting activity of CD95 in apoptosis-resistant tumor cells. (3) Glucose deprivation increased SNARK activity 3-fold in BHK fibroblasts. These findings identify SNARK as a glucose- and AICAriboside-regulated member of the AMPK-related gene family that represents a new candidate mediator of the cellular response to metabolic stress. (4)

References
1. Nagase, T.; et al., DNA Res. 5: 277-286, 1998.
2. Horike, N.; et al., J. Biol. Chem. 278: 18440-18447, 2003.
3. Legembre,P., et al., J. Biol. Chem. 279 (45), 46742-46747 (2004)
4. Lefebvre,D.L., et al., Biochem. J. 355 (PT 2), 297-305 (2001)