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Tubulin protein (>99% pure): porcine brain

Tubulin protein (>99% pure): porcine brain
$0.00
SKU
T240
Datasheet
MSDS
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Product Uses Include

  • IC50 & EC50 determinations for anti-tubulin ligands.
  • Microtubule binding studies
  • Tubulin monomer binding studies
  • HDAC6 studies
  • Microtubule activated kinesin ATPase assays
Click Here for Detailed Methods

Material
Tubulin protein has been purified from porcine brain by an adaptation of the method of Shelanski et al. (1), Further purification to >99% purity was achieved by cation exchange chromatography. Tubulin is supplied as a white lyophilized powder.

Fully active for polymerization, this product is lyophilized with a patented technology for increased stability and longevity. T240 is stable for 1 year at 4°C desiccated. If your project requires the same batch of tubulin for consistent results, it is highly recommended that the item is purchased in bulk in order to save time and money. This product can be used as a substitute for our highly purified bovine tubulin products (Cat. # TL238, T238 and T237) and behaves in an identical fashion.

Purity
Purity is determined by scanning densitometry of proteins on SDS-PAGE gels. Samples are >99% pure

t240

Figure 1: A 20 µg sample of T240 protein was separated by electrophoresis on a 10% SDS-PAGE gel and stained with Coomassie Blue. Protein quantitation was performed using the Precision Red Protein Assay Reagent (Cat. # ADV02).

Biological Activity
One unit of tubulin is defined as 5.0 mg of purified protein (as determined by the Precision Red Advanced Protein Assay Reagent cat. # ADV02). The biological activity of T240 is assessed by a tubulin polymerization assay. The ability of tubulin to polymerize into microtubules can be followed by observing an increase in optical density of the tubulin solution at 340 nm. A 5 mg/ml tubulin solution in General Tubulin Buffer buffer plus 5% glycerol and 1 mM GTP should achieve an OD340 nm reading between 0.75-1.10 in 30 min at 37°C when using a spectrophotometer pathlength of 0.8 cm (180 µl sample volume in a 1/2 area 96-well plate).

It should be noted that tubulin minus glycerol WILL NOT polymerize in G-PEM buffer until very high tubulin concentrations (>10 mg/ml). Even at these concentrations polymerization is comparatively slow. Efficient polymerization at low concentration of tubulin minus glycerol can be achieved by addition of a polymerization stimulating compound, e.g., glycerol, paclitaxel or DMSO.

References

Shelanski, M. L., et al. (1973). Proc. Natl. Acad. Sci. USA. 70, 765-768

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
Zhou, Qing et al.An orphan kinesin in Trypanosoma brucei regulates hook complex assembly and Golgi biogenesismBio2024
Babcock, N. S. et al.Ultraviolet Superradiance from Mega-Networks of Tryptophan in Biological ArchitecturesJournal of Physical Chemistry B2024
Benoit, Matthieu P.M.H. et al.Cryo-EM unveils kinesin KIF1A’s processivity mechanism and the impact of its pathogenic variant P305LNature Communications2024
Ballmer, Daniel et al.Dynamic localization of the chromosomal passenger complex in trypanosomes is controlled by the orphan kinesins KIN-A and KIN-BeLife2024
Lucero, Bobby et al.Design, Synthesis, and Evaluation of An Anti-trypanosomal [1,2,4]Triazolo[1,5-a]pyrimidine Probe for Photoaffinity Labeling StudiesChemMedChem2024
Singh, Sudhir K. et al.Noncanonical interaction with microtubules via the N-terminal nonmotor domain is critical for the functions of a bidirectional kinesinScience Advances2024
Heber, Simone et al.Tropomyosin 1-I/C coordinates kinesin-1 and dynein motors during oskar mRNA transportNature Structural & Molecular Biology 2024
Ceglowski, Julia et al.TTLL12 is required for primary ciliary axoneme formation in polarized epithelial cellsEMBO Reports2024
Fan, Yuanwei et al.A divergent tumor overexpressed gene domain and oligomerization contribute to SPIRAL2 function in stabilizing microtubule minus endsThe Plant Cell2024
Adler, Agnes et al.A structural and dynamic visualization of the interaction between MAP7 and microtubulesNature Communications2024
Liu, Xinglei et al.Kinesin-14 HSET and KlpA are non-processive microtubule motors with load-dependent power strokesNature Communications2024
Mahalingan, Kishore K. et al.Structural basis for α-tubulin-specific and modification state-dependent glutamylationNature Chemical Biology2024
Rai, Dipti et al.CAMSAPs and nucleation-promoting factors control microtubule release from γ-TuRCNature Cell Biology2024
Chauhan, Prashali et al.Ionic strength alters crosslinker-driven self-organization of microtubulesCytoskeleton2024
Yeh, Hung Wei et al.Cep57 regulates human centrosomes through multivalent interactionsProceedings of the National Academy of Sciences2024
Seo, Dahee et al.Poxvirus A51R proteins regulate microtubule stability and antagonize a cell-intrinsic antiviral responseCell Reports2024
Liang, Xiaolin et al.LncRNA TubAR complexes with TUBB4A and TUBA1A to promote microtubule assembly and maintain myelinationCell Discovery2024
Hu, Huiqing et al.A kinesin-13 family kinesin in Trypanosoma brucei regulates cytokinesis and cytoskeleton morphogenesis by promoting microtubule bundlingPLOS Pathogens2024
Tezuka, Toshiki et al.Clinical characteristics and pathophysiological properties of newly discovered LRRK2 variants associated with Parkinson's diseaseNeurobiology of Disease2024
Di Gregorio, Elisabetta et al.Raman Spectroscopy Reveals Photobiomodulation-Induced α-Helix to β-Sheet Transition in Tubulins: Potential Implications for Alzheimer’s and Other Neurodegenerative DiseasesNanomaterials2024
Fan, Weijing et al.A novel conductive microtubule hydrogel for electrical stimulation of chronic wounds based on biological electrical wiresJournal of Nanobiotechnology2024
Patrón, Lilian A. et al.Novel Brain-Penetrant, Small-Molecule Tubulin Destabilizers for the Treatment of GlioblastomaBiomedicines2024
Ranaivoson, Fanomezana Moutse et al.Nucleotide-free structures of KIF20A illuminate atypical mechanochemistry in this kinesin-6Open Biology2023
Gritsina, Galina et al.Chemokine receptor CXCR7 activates Aurora Kinase A and promotes neuroendocrine prostate cancer growthThe Journal of Clinical Investigation2023
Memarian, Fereshteh L. et al.Forming, Confining, and Observing Microtubule-Based Active NematicsJournal of Visualized Experiments (JoVE)2023
van den Berg, Cyntha M. et al.CSPP1 stabilizes growing microtubule ends and damaged lattices from the luminal sideJournal of Cell Biology2023
Wang, Yujuan et al.CCP5 and CCP6 retain CP110 and negatively regulate ciliogenesisBMC Biology2023
Bahri, Salima et al.1H-detected characterization of carbon–carbon networks in highly flexible protonated biomolecules using MAS NMRJournal of Biomolecular NMR2023
Minckley, Taylor F. et al.Zn2+ decoration of microtubules arrests axonal transport and displaces tau, doublecortin, and MAP2CJournal of Cell Biology2023
McGorty, Ryan J. et al.Kinesin and myosin motors compete to drive rich multiphase dynamics in programmable cytoskeletal compositesPNAS Nexus2023
Nithianantham, Stanley et al.The kinesin-5 tail and bipolar minifilament domains are the origin of its microtubule crosslinking and sliding activityMolecular biology of the cell2023
Hsieh, Yao Yu et al.Repositioning VU-0365114 as a novel microtubule-destabilizing agent for treating cancer and overcoming drug resistanceMolecular Oncology2023
Hoshino, Asumi et al.The microtubule-severing protein UNC-45A preferentially binds to curved microtubules and counteracts the microtubule-straightening effects of TaxolJournal of Biological Chemistry2023
Kalra, Aarat P. et al.Electronic Energy Migration in MicrotubulesACS Central Science2023
Hagita-Tatsumoto, Ayaka et al.Quantitative Microtubule Fractionation Technique to Separate Stable Microtubules, Labile Microtubules, and Free Tubulin in Mouse TissuesJoVE (Journal of Visualized Experiments)2023
McMillan, Pamela J. et al.Tau–RNA complexes inhibit microtubule polymerization and drive disease-relevant conformation changeBrain2023
Morishita, Jun et al.Identification of a small RhoA GTPase inhibitor effective in fission yeast and human cellsOpen Biology2023
Sharma, Sachin et al.Flavone-based dual PARP-Tubulin inhibitor manifesting efficacy against endometrial cancerJournal of Enzyme Inhibition and Medicinal Chemistry2023
Palumbo, Jacob et al.Directly Measuring Forces Within Reconstituted Active Microtubule BundlesJoVE (Journal of Visualized Experiments)2022
Henty-Ridilla, Jessica L.Visualizing Actin and Microtubule Coupling Dynamics In Vitro by Total Internal Reflection Fluorescence (TIRF) MicroscopyJoVE (Journal of Visualized Experiments)2022
Sasanpour, Mehrzad et al.Reconstituting and Characterizing Actin-Microtubule Composites with Tunable Motor-Driven Dynamics and MechanicsJoVE (Journal of Visualized Experiments)2022
Planelles-Herrero, Vicente Jose et al.Elongator stabilizes microtubules to control central spindle asymmetry and polarized trafficking of cell fate determinantsNature Cell Biology 2022
Feizabadi, Mitra Shojania et al.The Effect of Tau and Taxol on Polymerization of MCF7 Microtubules In VitroInternational Journal of Molecular Sciences2022
Yang, Yun et al.Altered succinylation of mitochondrial proteins, APP and tau in Alzheimer’s diseaseNature Communications2022
Kuzmić, Mira et al.Septin-microtubule association via a motif unique to isoform 1 of septin 9 tunes stress fibersJournal of Cell Science2022
Capizzi, Mariacristina et al.Developmental defects in Huntington's disease show that axonal growth and microtubule reorganization require NUMA1Neuron2022
Lee, Gloria et al.Myosin-driven actin-microtubule networks exhibit self-organized contractile dynamicsScience Advances2021
Hough, Cameron M. et al.Disassembly of microtubules by intense terahertz pulsesBiomedical Optics Express2021
Watson, Joseph L. et al.High-efficacy subcellular micropatterning of proteins using fibrinogen anchorsJournal of Cell Biology2021
Elahi, Montasir et al.High-fat diet–induced activation of SGK1 promotes Alzheimer’s disease–associated tau pathologyHuman Molecular Genetics2021
Zhou, Libin et al.Rare CASP6N73T variant associated with hippocampal volume exhibits decreased proteolytic activity, synaptic transmission defect, and neurodegenerationScientific Reports 2021
Budaitis, Breane G. et al.Pathogenic mutations in the kinesin-3 motor KIF1A diminish force generation and movement through allosteric mechanismsJournal of Cell Biology2021
Habicht, Juri et al.UNC-45A breaks the microtubule lattice independently of its effects on non-muscle myosin IIJournal of Cell Science2021
Gao, Li et al.A Robust, GFP-Orthogonal Photoswitchable Inhibitor Scaffold Extends Optical Control over the Microtubule CytoskeletonCell Chemical Biology2021
Nakayama, Shogo et al.Planar cell polarity induces local microtubule bundling for coordinated ciliary beatingJournal of Cell Biology2021
Ludzia, Patryk et al.Structural characterization of KKT4, an unconventional microtubule-binding kinetochore proteinStructure2021
Kundu, Tanushree et al.Coupling of dynamic microtubules to F-actin by Fmn2 regulates chemotaxis of neuronal growth conesJournal of Cell Science2021
Kaur, Simranpreet et al.Expansion of the phenotypic spectrum of de novo missense variants in kinesin family member 1A (KIF1A)Human Mutation2020
Baker, Stacey J. et al.A Contaminant Impurity, Not Rigosertib, Is a Tubulin Binding AgentMolecular Cell2020
Ricketts, Shea N. et al.Triggering Cation-Induced Contraction of Cytoskeleton Networks via MicrofluidicsFrontiers in Physics2020
Diwaker, Drishya et al.Deletion of the Pseudorabies virus gE/gI-US9p complex disrupts kinesin KIF1A and KIF5C recruitment during egress, and alters the properties of microtubule-dependent transport in vitroPLoS Pathogens2020
Ouyang, Changhan et al.Autophagic degradation of KAT2A/GCN5 promotes directional migration of vascular smooth muscle cells by reducing TUBA/α-tubulin acetylationAutophagy2020
Müller-Deku, Adrian et al.Photoswitchable pac*******-based microtubule stabilisers allow optical control over the microtubule cytoskeletonNature Communications2020
Alatrash, Nagham et al.Disruption of microtubule function in cultured human cells by a cytotoxic ruthenium(ii) polypyridyl complexChemical Science2020
Kraus, Yvonne et al.Isoquinoline-based biaryls as a robust scaffold for microtubule inhibitorsEuropean Journal of Medicinal Chemistry2020
Aher, Amol et al.CLASP Mediates Microtubule Repair by Restricting Lattice Damage and Regulating Tubulin IncorporationCurrent Biology2020
Gunji, Shizuka et al.Excess Pyrophosphate Restrains Pavement Cell Morphogenesis and Alters Organ Flatness in Arabidopsis thalianaFrontiers in Plant Science2020
Francis, Madison L. et al.Non-monotonic dependence of stiffness on actin crosslinking in cytoskeleton compositesSoft Matter2019
Leong, Su Ling et al.Reconstitution of Microtubule Nucleation In Vitro Reveals Novel Roles for Mzt1Current Biology2019
Lopes, Joseph et al.Membrane mediated motor kinetics in microtubule gliding assaysScientific Reports2019
Nakos, Konstantinos et al.Regulation of microtubule plus end dynamics by septin 9Cytoskeleton2019
Ng, Cai Tong et al.Electron cryotomography analysis of Dam1C/DASH at the kinetochore-spindle interface in situJournal of Cell Biology2019
Guzman-Sepulveda, Jose Rafael et al.Tubulin Polarizability in Aqueous SuspensionsACS Omega2019
Karasmanis, Eva P. et al.Erratum: Polarity of Neuronal Membrane Traffic Requires Sorting of Kinesin Motor Cargo during Entry into Dendrites by a Microtubule-Associated Septin (Developmental Cell (2018) 46(2) (204–218.e7), (S1534580718304982) (10.1016/j.devcel.2018.06.013))Developmental Cell2018
Ganguly, Anindya et al.Importin-β Directly Regulates the Motor Activity and Turnover of a Kinesin-4Developmental Cell2018
Fan, Yuanwei et al.The Arabidopsis SPIRAL2 Protein Targets and Stabilizes Microtubule Minus EndsCurrent Biology2018
Romé, Pierre et al.A novel microtubule nucleation pathway for meiotic spindle assembly in oocytesJournal of Cell Biology2018
Tripathy, Ratna et al.Mutations in MAST1 Cause Mega-Corpus-Callosum Syndrome with Cerebellar Hypoplasia and Cortical MalformationsNeuron2018
Murray, John W. et al.Reduction of organelle motility by removal of potassium and other solutesPLoS ONE2017
Kandel, Mikhail E. et al.Label-Free Imaging of Single Microtubule Dynamics Using Spatial Light Interference MicroscopyACS Nano2017
Okeyoshi, Kosuke et al.Methods for the self-integration of megamolecular biopolymers on the drying air-LC interfaceJournal of Visualized Experiments2017
Majellaro, Maria et al.Investigating Structural Requirements for the Antiproliferative Activity of Biphenyl NicotinamidesChemMedChem2017
Onder, Seda et al.Monoclonal Antibody That Recognizes Diethoxyphosphotyrosine-Modified Proteins and Peptides Independent of Surrounding Amino AcidsChemical Research in Toxicology2017
Monda, Julie K. et al.Microtubule Tip Tracking by the Spindle and Kinetochore Protein Ska1 Requires Diverse Tubulin-Interacting SurfacesCurrent Biology2017
Howes, Stuart C. et al.Structural differences between yeast and mammalian microtubules revealed by cryo-EMJournal of Cell Biology2017
Ghaly, Peter E. et al.A new antiproliferative noscapine analogue: Chemical synthesis and biological evaluationOncotarget2016
Lor, Chai et al.A simple experimental model to investigate force range for membrane nanotube formationFrontiers in Materials2016
Ayoub, Ahmed Taha et al.Antitumor Activity of Lankacidin Group Antibiotics Is Due to Microtubule Stabilization via a Pac*******-like MechanismJournal of Medicinal Chemistry2016
Matsuyama, Tomonori et al.Midaz(olam) inhibits the hypoxia-induced up-regulation of erythropoietin in the central nervous systemEuropean Journal of Pharmacology2015
Hara, Yuki et al.Dynein-Based Accumulation of Membranes Regulates Nuclear Expansion in Xenopus laevis Egg ExtractsDevelopmental Cell2015
Kim, Kyongwan et al.Electric field-induced reversible trapping of microtubules along metallic glass microwire electrodesJournal of Applied Physics2015
Taberner, Núria et al.Reconstituting Functional Microtubule-Barrier InteractionsMethods in Cell Biology2014
Hosono, Hitomi et al.The murine Gcap14 gene encodes a novel microtubule binding and bundling proteinFEBS letters2012
Kawakami, Fumitaka et al.LRRK2 phosphorylates tubulin-associated tau but not the free molecule: LRRK2-mediated regulation of the tau-tubulin association and neurite outgrowthPloS one2012
Berezniuk, Iryna et al.Cytosolic Carboxypeptidase 1 Is Involved in Processing α- and β-TubulinThe Journal of Biological Chemistry2012
Wu, Hui Yuan et al.A structural and functional analysis of Nna1 in Purkinje cell degeneration (pcd) miceThe FASEB Journal2012
Wells, David B. et al.Mechanical properties of a complete microtubule revealed through molecular dynamics simulationBiophysical Journal2010
Iuchi, Katsuya et al.heterocyclic organobismuth(III) compound targets tubulin to induce G 2/M arrest in hela cellsJournal of Pharmacological Sciences2009
Ligon, Lee A. et al.The Microtubule Plus-End Proteins EB1 and Dynactin Have Differential Effects on Microtubule PolymerizationMolecular Biology of the Cell2003
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Question 1:  What is the proper way to store the tubulin to insure maximum stability and activity?

Answer 1:  The recommended storage condition for the lyophilized tubulin product is 4°C with desiccant to maintain humidity at <10% humidity.  Under these conditions the protein is stable for 6 months.  Lyophilized protein can also be stored desiccated at -70°C where it will be stable for 6 months.  However, at -70°C the rubber seal in the lid of the tube could crack and allow in moisture.  Therefore we recommend storing at 4°C.  If stored at -70°C, it is imperative to include desiccant with the lyophilized protein if this storage condition is utilized.  After reconstituting the protein as directed, the concentrated protein in G-PEM buffer should be aliquoted, snap frozen in liquid nitrogen and stored at -70°C (stable for 6 months).  NOTE: It is very important to snap freeze the tubulin in liquid nitrogen as other methods of freezing will result in significantly reduced activity.  Defrost rapidly by placing in a room temperature water bath for 1 min.  Avoid repeated freeze/thaw cycles.

 

Question 2:  Why does Cytoskeleton recommend the use of general tubulin buffer and GTP for resuspending tubulin?

Answer 2:  We recommend resuspending tubulin in general tubulin buffer + GTP to maintain tubulin monomer protein stability and conformation and to provide the necessary components for polymerization.  For resuspension, we recommend using a general tubulin buffer (Cat. # BST01-001) which consists of 80 mM PIPES, 2 mM MgCl2, 1 mM EGTA, pH 7.0, supplemented with 1 mM GTP (Cat. # BST06-001).  Tubulin requires GTP and magnesium ions for proper stability and conformation, even in its monomeric state.  GTP is also required for the polymerization process as its hydrolysis during tubulin polymerization is necessary for polymerization to occur.  EGTA is a chelator of calcium which is a potent inhibitor of tubulin polymerization.  Glycerol is often added to a final concentration of 5 - 10% to enhance polymerization; however, glycerol is not necessary for the maintenance of biologically active tubulin and does not need to be included when reconstituting and storing tubulin.  When aliquoting reconstituted tubulin for storage, it is essential to aliquot and snap-freeze tubulin in liquid nitrogen at a concentration of >6 mg/ml to preserve tubulin’s biological activity.  Then the aliquots should be stored at -70°C.  When thawing the aliquots, thaw rapidly in a room temperature water bath and place on ice until right before experimental use.

 

Question 3: How does porcine tubulin compare to bovine tubulin?

Answer 3: Click here for an in-depth comparison of porcine and bovine tubulin.

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

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