Phospho-(Ser/Thr) AMPK Substra

5759:

ELISA Peptide , Immunoprecipitation , Western Blotting

Phospho-(Ser/Thr) AMPK Substrate (P-S/T² -102) Rabbit mAb preferentially recognizes endogenous proteins and peptides bearing the LXRXXpS/pT motif. The antibody also cross-reacts with proteins and peptides that only harbor an RXXpS/pT motif.

Monoclonal antibody is produced by immunizing rabbits with a synthetic LXRXX(S*/T*) peptide library. The antibody is formulated from two rabbit monoclones in order to cover a broad range of reactivity.

Western Blotting

Western blot analysis of extracts from various mouse tissues using Phospho-(Ser/Thr) AMPK Substrate (P-S/T² -102) Rabbit mAb.

Western Blotting

Western blot analysis of extracts from H1650 cells, untreated or treated with phenformin (5 mM, 1 hr), using Phospho-(Ser/Thr) AMPK Substrate (P-S/T² -102) Rabbit mAb. Western blot was imaged using Odyssey^® Infrared Imaging System (LI-COR^® Biosciences).

IP

Immunoprecipitation of extracts from H1650 cells using Phospho-(Ser/Thr) AMPK Substrate (P-S/T² -102) Rabbit mAb. Lane 1 shows 10% input. Western blot analysis was performed using the same antibody.

AMP-activated protein kinase (AMPK) is highly conserved from yeast to plants and animals and plays a key role in the regulation of energy homeostasis (1). AMPK is a heterotrimeric complex composed of a catalytic α subunit and regulatory β and γ subunits, each of which is encoded by two or three distinct genes (α1, 2; β1, 2; γ1, 2, 3) (2). The kinase is activated by an elevated AMP/ATP ratio due to cellular and environmental stress, such as heat shock, hypoxia, and ischemia (1). The tumor suppressor LKB1, in association with accessory proteins STRAD and MO25, phosphorylates AMPKα at Thr172 in the activation loop, and this phosphorylation is required for AMPK activation (3-5). AMPKα is also phosphorylated at Thr258 and Ser485 (for α1; Ser491 for α2). The upstream kinase and the biological significance of these phosphorylation events have yet to be elucidated (6). The β1 subunit is post-translationally modified by myristoylation and multi-site phosphorylation including Ser24/25, Ser96, Ser101, Ser108, and Ser182 (6,7). Phosphorylation at Ser108 of the β1 subunit seems to be required for the activation of AMPK enzyme, while phosphorylation at Ser24/25 and Ser182 affects AMPK localization (7). Several mutations in AMPKγ subunits have been identified, most of which are located in the putative AMP/ATP binding sites (CBS or Bateman domains). Mutations at these sites lead to reduction of AMPK activity and cause glycogen accumulation in heart or skeletal muscle (1,2). Accumulating evidence indicates that AMPK not only regulates the metabolism of fatty acids and glycogen, but also modulates protein synthesis and cell growth through EF2 and TSC2/mTOR pathways, as well as blood flow via eNOS/nNOS (1).

AMPK phosphorylates consensus motif (L/M)XRXX(S/T)XXXL (8). Antibodies recognizing the LXRXX(S/T) motif are very useful in the identification of AMPK substrates.

1. Hardie, D.G. (2004) J Cell Sci 117, 5479-87.
2. Carling, D. (2004) Trends Biochem Sci 29, 18-24.
3. Hawley, S.A. et al. (1996) J Biol Chem 271, 27879-87.
4. Lizcano, J.M. et al. (2004) EMBO J 23, 833-43.
5. Shaw, R.J. et al. (2004) Proc Natl Acad Sci USA 101, 3329-35.
6. Woods, A. et al. (2003) J Biol Chem 278, 28434-42.
7. Warden, S.M. et al. (2001) Biochem J 354, 275-83.
8. Gwinn, D.M. et al. (2008) Mol Cell 30, 214-26.

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• 9812 KinomeView^® Profiling Kit
• 5760 PTMScan^® Phospho-AMPK Substrate Motif (LXRXXS*/T*) Immunoaffinity Beads
• 5564 PTMScan^® Phospho-AMPK Substrate Motif (LXRXXS*/T*) Kit
• 9957 AMPK and ACC Antibody Sampler Kit
• 9839 AMPK Subunit Antibody Sampler Kit
• 8208 PhosphoPlus^® AMPKα (Thr172) Antibody Duet
• 6620 SignalSilence^® AMPKα2 siRNA I
• 6630 SignalSilence^® AMPKα2 siRNA II

For Research Use Only. Not For Use In Diagnostic Procedures.