Recombinant Human MCAK Protein: Properties & Research Uses

Recombinant Human MCAK Protein is a kinesin-13 family microtubule depolymerizing enzyme that plays a critical role in mitotic spindle assembly, chromosome segregation, and microtubule dynamics, making it an essential reagent for cytoskeleton and cell division research.

Gene Name

KIF2C (also known as MCAK)

Protein Family

Kinesin-13 superfamily (microtubule depolymerizing kinesin)

Primary Function

ATP-dependent microtubule depolymerization at plus and minus ends

Biological Role

Regulates mitotic spindle dynamics and accurate chromosome segregation

SKU

rp375162

Source / Expression

Recombinant protein; expression system details to be confirmed with product datasheet

Research Applications

In vitro microtubule depolymerization assays, ATPase activity assays, cell cycle studies, protein interaction studies

What Is MCAK? Gene, Structure, and Protein Family

Mitotic centromere-associated kinesin (MCAK), encoded by the gene KIF2C, is a member of the kinesin-13 superfamily of motor proteins. Unlike conventional kinesins that use ATP hydrolysis to walk along microtubule tracks, MCAK and other kinesin-13 family members do not translocate processively. Instead, they harness ATP hydrolysis to catalyze the removal of tubulin dimers from both the plus and minus ends of microtubules, effectively depolymerizing the polymer lattice.

Structurally, MCAK contains an N-terminal regulatory domain, a central motor domain responsible for ATP hydrolysis and microtubule binding, and a C-terminal domain that also contributes to microtubule interaction. The protein functions as a dimer in its active state. Regulation of MCAK activity occurs through phosphorylation by several mitotic kinases, including Aurora B, which is a well-established regulatory mechanism during cell division.

Recombinant Human MCAK Protein (rp375162) enables researchers to study these biochemical properties in controlled in vitro settings, providing a defined and reproducible reagent for mechanistic investigations.

Biological Functions of MCAK in Cell Division

MCAK plays indispensable roles in ensuring the fidelity of cell division. Its primary function is to regulate microtubule dynamics within the mitotic spindle. By depolymerizing microtubules at kinetochores and spindle poles, MCAK contributes to the correction of erroneous kinetochore–microtubule attachments, such as syntelic and merotelic attachments, which if left uncorrected would lead to chromosome mis-segregation and aneuploidy.

Key biological functions attributed to MCAK include:

• Kinetochore error correction: MCAK, activated by Aurora B kinase at the inner centromere, destabilizes incorrect kinetochore–microtubule attachments and promotes their replacement with correct amphitelic attachments.
• Spindle assembly and pole focusing: MCAK activity at spindle poles helps maintain proper spindle morphology and pole integrity during mitosis.
• Microtubule flux: MCAK contributes to poleward microtubule flux by depolymerizing microtubule minus ends at spindle poles.
• Cytokinesis: MCAK has been implicated in regulating microtubule organization during the later stages of cell division.

Dysregulation of MCAK expression or activity has been associated with chromosomal instability and has been observed in various cancer types, making it a subject of interest in oncology research.

Research Applications of Recombinant Human MCAK Protein

Recombinant Human MCAK Protein (rp375162) is a versatile reagent supporting a range of biochemical and cell biology experiments. Key research applications include:

• In vitro microtubule depolymerization assays: Recombinant MCAK can be used in reconstituted assays with purified tubulin to quantitatively measure microtubule depolymerization activity, determine kinetic parameters, and study the effect of inhibitors or regulatory proteins.
• ATPase activity assays: The intrinsic ATPase activity of MCAK, which is stimulated by microtubules, can be measured using colorimetric or fluorometric phosphate release assays to characterize inhibitors or activators.
• Protein–protein interaction studies: Recombinant MCAK serves as bait or prey in pull-down assays, co-immunoprecipitation experiments, and surface plasmon resonance (SPR) studies to identify and characterize binding partners.
• Structural biology: Purified recombinant MCAK can be used in crystallography or cryo-electron microscopy studies to elucidate the molecular mechanism of microtubule depolymerization.
• Biochemical characterization of MCAK mutations: Disease- or cancer-associated variants of MCAK can be compared with wild-type recombinant protein to understand functional consequences.
• Drug discovery and screening: MCAK is a potential anticancer target; recombinant protein enables screening of small-molecule inhibitor libraries in enzymatic assays.

Researchers are advised to consult the product datasheet for specific activity values, recommended storage conditions, and buffer compositions before designing experiments.

Handling, Storage, and Quality Considerations

To maintain the activity and integrity of Recombinant Human MCAK Protein, researchers should follow standard best practices for handling recombinant proteins:

• Storage: Store the protein according to the conditions specified in the product datasheet. Recombinant proteins are generally stored at −80 °C for long-term stability. Avoid repeated freeze–thaw cycles, which can lead to aggregation and loss of activity.
• Aliquoting: Upon first thaw, prepare single-use aliquots to minimize freeze–thaw degradation.
• Buffer compatibility: Ensure the protein storage buffer is compatible with your downstream assay conditions. If buffer exchange is required, use desalting columns or dialysis.
• Protein concentration: Verify protein concentration using an appropriate assay (e.g., BCA or Bradford assay) prior to use, as shipping and freeze–thaw steps may affect the stated concentration.
• Activity verification: For critical experiments, confirm depolymerization or ATPase activity using an orthogonal assay before proceeding with primary experiments.

All handling conditions, lot-specific quality control data, and purity information are provided in the certificate of analysis (CoA) accompanying each product lot. Contact Aladdin Scientific technical support for further guidance.

Frequently asked questions

What is the difference between MCAK and other kinesin motor proteins?

Unlike conventional kinesin-1 family members that move processively along microtubules to transport cargo, MCAK belongs to the kinesin-13 family and functions as a microtubule depolymerase. It uses ATP hydrolysis to remove tubulin dimers from microtubule ends rather than to generate unidirectional movement, and it acts at both plus and minus ends of microtubules.

How is MCAK activity regulated in cells?

MCAK activity is primarily regulated by phosphorylation. Aurora B kinase, concentrated at the inner centromere during mitosis, phosphorylates MCAK to stimulate its depolymerizing activity at kinetochores and to promote error correction of improper kinetochore–microtubule attachments. Other kinases and regulatory proteins also modulate MCAK activity in a cell-cycle-dependent manner.

What assay formats are suitable for testing Recombinant Human MCAK Protein activity?

The two most common formats are in vitro microtubule depolymerization assays—where polymerized microtubules (often fluorescently labeled or stabilized with taxol) are incubated with MCAK in the presence of ATP and depolymerization is monitored by sedimentation or fluorescence microscopy—and malachite green ATPase assays, which quantify inorganic phosphate released during ATP hydrolysis. Researchers should refer to the product datasheet for recommended assay conditions.

Is Recombinant Human MCAK Protein suitable for antibody production or immunization?

Recombinant proteins can in principle be used as antigens for antibody production; however, the specific suitability of this product for immunization depends on its purity, the presence of tags, and the folding state of the protein. Researchers intending to use rp375162 for antigen production should contact Aladdin Scientific technical support to confirm lot-specific details and purity data before proceeding.

Why is MCAK relevant to cancer research?

MCAK is implicated in chromosomal instability (CIN), a hallmark of many cancers characterized by ongoing errors in chromosome segregation. Overexpression or dysregulation of MCAK has been reported in several tumor types, and its role in kinetochore error correction makes it an attractive target for therapeutic intervention. Recombinant MCAK protein facilitates biochemical screening of inhibitor candidates.

References

1. Walczak CE, Mitchison TJ, Desai A. XKCM1: a Xenopus kinesin-related protein that regulates microtubule dynamics during mitotic spindle assembly. Cell. 1996;84(1):37–47.
2. Desai A, Verma S, Mitchison TJ, Walczak CE. Kin I kinesins are microtubule-destabilizing enzymes. Cell. 1999;96(1):69–78.
3. Andrews PD, Ovechkina Y, Morrice N, et al. Aurora B regulates MCAK at the mitotic centromere. Dev Cell. 2004;6(2):253–268.
4. Wordeman L. How kinesin motor proteins drive mitotic spindle function: lessons from molecular assays. Semin Cell Dev Biol. 2010;21(3):260–268.

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.