Kinesin ATPase Assay Services
Target-specific Assays

Kinesin ATPase Assay Services

Kinesin superfamily proteins (KIFs) are a large family of molecular motor proteins, that share a highly conserved motor domain. Molecular motors convert chemical energy into mechanical work to achieve active cellular processes. Kinesins move along microtubule filaments utilizing energy from ATP hydrolysis and play important role in cell division and transport of vesicles and organelles within cells.

Altered expressions of several kinesins are shown in various cancers, fueling the aggressive nature of cancers leading to genomic instability. Eg5/KIF11 overexpression is shown in various cancers including hepatic carcinoma, lung, prostate, colorectal, gastric, and pancreatic cancers. Small molecule inhibitors of KIF11/Eg5 and CENP-E/ KIF10 are shown to result in mitotic arrest leading to apoptosis in tumor cell lines. Recently, an inhibitor targeting Kinesin 8 family member KIF18A, has entered clinical trial for treatment of solid tumors with high chromosomal instability several other kinesins such MCAK, MKLP2/KIF20A, KIF22 and MKLP1/KIF23 are shown to be aberrantly regulated in several cancers attracted interest in anti-cancer therapeutic discovery.

The Kinesin Superfamily

Kinesin superfamily contains 46 kinesins classified into 14 subfamilies based on their structural features.

Reference:
https://doi.org/10.1073/pnas.111145398

Kinesin Tree

Kinesin ATPase Assay Details

  • Assay Format
  • Assay Setup
Assay Format

Kinesin ATPase Assay Format

Reaction Biology offers ATPase activity inhibitor screening assays for kinesin targets via Promega’s ADP GloTM detection system.

In the presence of stabilized microtubules, kinesin enzyme activity utilizes ATP, producing ADP and Pi. ADP formation is quantified to measure kinesin enzymatic activity. In the presence of compounds inhibiting kinesin, ATPase activity formation of ADP is lowered compared to that of no inhibitor/vehicle control.

Assay Setup

Kinesin ATPase Assay Setup

Setups: Single-dose screening in duplicates or IC50 value determination with 5 or 10 concentrations. Other screening formats are available upon request.

Controls: No inhibitor (DMSO vehicle) control and for every assay, one target-specific control compound is tested in 10-dose IC50 format.

Turnaround time: 10 business days for standard projects. Expedited scheduling and data delivery can be arranged prior to the commencement of the studies.

Report: The raw data (ADP product formation), % enzyme activity relative to control and control compound IC50 values will be reported in Excel format for single-dose assays. For IC50 orders, raw data (ADP product formation), % enzyme activity relative to control, IC50 values, and curve fitting will be delivered in Excel format. Assay conditions, target, and substrate information are available upon request. Requirements for this information should be noted prior to the commencement of the study.

Screening facility: This assay is performed at our screening facility in Malvern, PA, US.

Compound requirements: In brief, for a standard project, 20ul of a 10mM DMSO stock or solid material is needed. Less material is needed for large scale screening. Please refer to our FAQs for information regarding compound preparation and shipping.