Product Uses Include
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.
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).
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.
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.
|Lin, Ying H. et al.||Site-specific acetyl-mimetic modification of cardiac troponin I modulates myofilament relaxation and calcium sensitivity||Journal of Molecular and Cellular Cardiology||2020||ISSN 1095-8584|
|Schmidt, William et al.||Lysine acetylation of F-actin decreases tropomyosin-based inhibition of actomyosin activity||The Journal of biological chemistry||2020||ISSN 1083--351X|
|Yamada, Yurika et al.||Cardiac muscle thin filament structures reveal calcium regulatory mechanism||Nature Communications||2020||ISSN 2041-1723|
|Seervai, Riyad N.H. et al.||The Huntingtin-interacting protein SETD2/HYPB is an actin lysine methyltransferase||Science Advances||2020||ISSN 2375-2548|
|Ravichandran, Akshaya et al.||A novel actin binding drug with in vivo efficacy||Antimicrobial Agents and Chemotherapy||2019||ISSN 1098-6596|
|Gu, Weihong et al.||Mono-fullerenols modulating cell stiffness by perturbing actin bundling||Nanoscale||2018||ISSN 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 cells||Biomedicine and Pharmacotherapy||2018||ISSN 1950-6007|
|Gu, Weihong et al.||Highly Dispersed Fullerenols Hamper Osteoclast Ruffled Border Formation by Perturbing Ca2+ Bundles||Small||2018||ISSN 1613-6829|
|Cui, Jin et al.||Leptolyngbyolides, Cytotoxic Macrolides from the Marine Cyanobacterium Leptolyngbya sp.: Isolation, Biological Activity, and Catalytic Asymmetric Total Synthesis||Chemistry - A European Journal||2017||ISSN 1521-3765|
|Lin, Yi et al.||Toxic PR Poly-Dipeptides Encoded by the C9orf72 Repeat Expansion Target LC Domain Polymers||Cell||2016||ISSN 1097-4172|
|Brody, Matthew J. et al.||LRRC10 is required to maintain cardiac function in response to pressure overload||American Journal of Physiology - Heart and Circulatory Physiology||2016||ISSN 1522-1539|
|Lundquist, Mark R. et al.||Redox modification of nuclear actin by MICAL-2 regulates SRF signaling||Cell||2014||ISSN 1097-4172|
|Wang, Y et al.||Fluorescence imaging with one-nanometer accuracy (FIONA)||JoVE (Journal of …||2014||Article Link|
|Mohammad, Ibrahim et al.||Flightless I is a focal adhesion-associated actin-capping protein that regulates cell migration||The FASEB Journal||2012||ISSN 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.)||2012||ISSN 1528-3658|
|Mohammad, Ibrahim et al.||Flightless I is a focal adhesion‐associated actin‐capping protein that regulates cell migration||The FASEB Journal||2012||ISSN 0892--6638|
|Brody, Matthew J. et al.||Ablation of the Cardiac-Specific Gene Leucine-Rich Repeat Containing 10 (Lrrc10) Results in Dilated Cardiomyopathy||PLoS ONE||2012||ISSN 1932-6203|
|Windhorst, Sabine et al.||Functional role of inositol-1,4,5-trisphosphate-3-kinase-A for motility of malignant transformed cells||International Journal of Cancer||2011||ISSN 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 adhesion||Cell Communication and Adhesion||2006||ISSN 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 membrane||PLoS Biology||2004||ISSN 1544-9173|
|Ishikawa, Takashi et al.||Subdomain organization of the Acanthamoeba myosin IC tail from cryo-electron microscopy||Proceedings of the National Academy of Sciences of the United States of America||2004||ISSN 0027-8424|
|Arora, Pamela D. et al.||Gelsolin Mediates Collagen Phagocytosis through a Rac-dependent Step||Molecular Biology of the Cell||2004||ISSN 1059-1524|
|Upadhyaya, Arpita et al.||Probing polymerization forces by using actin-propelled lipid vesicles||Proceedings of the National Academy of Sciences of the United States of America||2003||ISSN 0027-8424|
|Balcer, Heath I. et al.||No Title||2003||PMID 14680631|
|Loomis, Patricia A. et al.||Espin cross-links cause the elongation of microvillus-type parallel actin bundles in vivo||2003||PMID 14657236|
|Humphries, Christine L. et al.||Direct regulation of Arp2/3 complex activity and function by the actin binding protein coronin||2002||ISSN 0021--9525|
|Engqvist-Goldstein, Åsa E.Y. et al.||The actin-binding protein Hip1R associates with clathrin during early stages of endocytosis and promotes clathrin assembly in vitro||Journal of Cell Biology||2001||ISSN 0021-9525|
|Tamura, Minoru et al.||Deactivation of neutrophil NADPH oxidase by actin-depolymerizing agents in a cell-free system||Biochemical Journal||2000||ISSN 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.
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