Actin protein (>95% pure): rabbit skeletal muscle

Actin protein (>95% pure): rabbit skeletal muscle
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Product Uses Include

  • Identification and characterization of muscle actin binding proteins
  • In vitro actin polymerization studies
  • Antibody standard for Western blot analysis

Material
Actin protein has been purified from rabbit skeletal muscle. AKL99 actin is greater than 99% pure and AKL95 is greater than 95% pure. Muscle actin has an approximate molecular weight of 43 kDa. Rabbit muscle actin is supplied as a white lyophilized powder. The lyophilized protein when stored desiccated to < 10% humidity at 4°C is stable for 6 months. When re constituted in distilled water to 10 mg/ml, the protein is in the following buffer: 5 mM Tris-HCl pH 8.0, 0.2 mM CaCl2, 0.2 mM ATP, 5% sucrose, and 1% dextran.

Purity
Protein purity is determined by scanning densitometry of Coomassie Blue stained protein on a 12% polyacrylamide gel. AKL99 consists of >99% pure muscle actin while AKL95 is >95% pure (see Figure 1).

aklgel

Figure 1: Figure 1. Purities of rabbit skeletal muscle actin protein. 100 µg of >99% pure (AKL99) and >95% pure (AKL95) rabbit skeletal muscle actin were run on SDS-PAGE gels and stained with coomassie blue. The arrow indicates actin protein, the arrowhead an α-actinin contaminant (115 kDa). The minor impurities in the purified actins are predominantly actin binding proteins such as α-actinin and gelsolin.

Biological Activity
The biological activity of muscle actinis determined by its ability to efficiently polymerize into filaments (F-actin) in vitro and separate from unpolymerized components in a spin down assay. Stringent quality control ensures that AKL99 produces > 90% F-actin and AKL95 produces > 80% F-actin in this assay.

For product Datasheets and MSDSs please click on the PDF links below.   For additional information, click on the FAQs tab above or contact our Technical Support department at tservice@cytoskeleton.com

AuthorTitleJournalYearArticle Link
Lin, Ying H. et al.Site-specific acetyl-mimetic modification of cardiac troponin I modulates myofilament relaxation and calcium sensitivityJournal of Molecular and Cellular Cardiology2020ISSN 1095-8584
Schmidt, William et al.Lysine acetylation of F-actin decreases tropomyosin-based inhibition of actomyosin activityThe Journal of biological chemistry2020ISSN 1083--351X
Yamada, Yurika et al.Cardiac muscle thin filament structures reveal calcium regulatory mechanismNature Communications2020ISSN 2041-1723
Seervai, Riyad N.H. et al.The Huntingtin-interacting protein SETD2/HYPB is an actin lysine methyltransferaseScience Advances2020ISSN 2375-2548
Ravichandran, Akshaya et al.A novel actin binding drug with in vivo efficacyAntimicrobial Agents and Chemotherapy2019ISSN 1098-6596
Gu, Weihong et al.Mono-fullerenols modulating cell stiffness by perturbing actin bundlingNanoscale2018ISSN 2040--3372
Bertier, Laurence et al.Nanobodies targeting cortactin proline rich, helical and actin binding regions downregulate invadopodium formation and matrix degradation in SCC-61 cancer cellsBiomedicine and Pharmacotherapy2018ISSN 1950-6007
Gu, Weihong et al.Highly Dispersed Fullerenols Hamper Osteoclast Ruffled Border Formation by Perturbing Ca2+ BundlesSmall2018ISSN 1613-6829
Cui, Jin et al.Leptolyngbyolides, Cytotoxic Macrolides from the Marine Cyanobacterium Leptolyngbya sp.: Isolation, Biological Activity, and Catalytic Asymmetric Total SynthesisChemistry - A European Journal2017ISSN 1521-3765
Lin, Yi et al.Toxic PR Poly-Dipeptides Encoded by the C9orf72 Repeat Expansion Target LC Domain PolymersCell2016ISSN 1097-4172
Brody, Matthew J. et al.LRRC10 is required to maintain cardiac function in response to pressure overloadAmerican Journal of Physiology - Heart and Circulatory Physiology2016ISSN 1522-1539
Lundquist, Mark R. et al.Redox modification of nuclear actin by MICAL-2 regulates SRF signalingCell2014ISSN 1097-4172
Wang, Y et al.Fluorescence imaging with one-nanometer accuracy (FIONA)JoVE (Journal of …2014Article Link
Mohammad, Ibrahim et al.Flightless I is a focal adhesion-associated actin-capping protein that regulates cell migrationThe FASEB Journal2012ISSN 1530--6860
Reay, Daniel P. et al.Effect of nuclear factor κB inhibition on serotype 9 adeno-associated viral (AAV9) minidystrophin gene transfer to the mdx mouse.Molecular medicine (Cambridge, Mass.)2012ISSN 1528-3658
Mohammad, Ibrahim et al.Flightless I is a focal adhesion‐associated actin‐capping protein that regulates cell migrationThe FASEB Journal2012ISSN 0892--6638
Brody, Matthew J. et al.Ablation of the Cardiac-Specific Gene Leucine-Rich Repeat Containing 10 (Lrrc10) Results in Dilated CardiomyopathyPLoS ONE2012ISSN 1932-6203
Windhorst, Sabine et al.Functional role of inositol-1,4,5-trisphosphate-3-kinase-A for motility of malignant transformed cellsInternational Journal of Cancer2011ISSN 1097--0215
Pappas, Derek J. et al.Direct interaction of the C-terminal domain of α-catenin and F-actin is necessary for stabilized cell-cell adhesionCell Communication and Adhesion2006ISSN 1541-9061
Holaska, James M. et al.Emerin caps the pointed end of actin filaments: Evidence for an actin cortical network at the nuclear inner membranePLoS Biology2004ISSN 1544-9173
Ishikawa, Takashi et al.Subdomain organization of the Acanthamoeba myosin IC tail from cryo-electron microscopyProceedings of the National Academy of Sciences of the United States of America2004ISSN 0027-8424
Arora, Pamela D. et al.Gelsolin Mediates Collagen Phagocytosis through a Rac-dependent StepMolecular Biology of the Cell2004ISSN 1059-1524
Upadhyaya, Arpita et al.Probing polymerization forces by using actin-propelled lipid vesiclesProceedings of the National Academy of Sciences of the United States of America2003ISSN 0027-8424
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Loomis, Patricia A. et al.Espin cross-links cause the elongation of microvillus-type parallel actin bundles in vivo2003PMID 14657236
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Tamura, Minoru et al.Deactivation of neutrophil NADPH oxidase by actin-depolymerizing agents in a cell-free systemBiochemical Journal2000ISSN 0264-6021

Question 1:  What is the best way to store actin proteins to insure maximum stability and shelf-life?

Answer 1:  Cytoskeleton provides all of our actin proteins as lyophilized powders so that they can be shipped at room temperature.  Upon receipt, the lyophilized powders should be stored at 4°C in a sealed container with desiccant.  It is important to monitor the freshness of the desiccant and insure that it continues to absorb moisture to protect the lyophilized actins.  With proper storage, the lyophilized actins are guaranteed to be stable for 6 months from the date of purchase.  Alternatively, actins can be immediately resuspended at the concentration recommended, aliquoted, snap-frozen in liquid nitrogen and stored at -70°C.  The frozen aliquots will be stable for 6 months.  When thawing frozen aliquots, it is important to thaw in a room temperature water bath.

 

Question 2:  What is the best way to store F-actin after polymerizing?

Answer 2:  G-actin is stable for two days at 4°C and requires a divalent cation, pH 6.5 - 8.0 and ATP for stability.  F-actin is stable and can be stored at 4°C for 1-2 weeks.  F-actin requires ATP (0.2 mM) and Mg2+ (2 mM) for stability and is unstable below pH 6.5 and above pH 8.5.  F-actin is not stable to freezing.  F-actin can be transferred to a variety of buffers (e.g. HEPES, phosphate, etc) without detrimental effects.  We recommend the addition of antibacterial agents such as 100 μg/ml ampicillin and 10 μg/ml chloramphenicol when storing F-actin at 4°C.

 

If you have any questions concerning this product, please contact our Technical Service department at tservice@cytoskeleton.com.