Laboratory of Motor Proteins

Head: Andrzej A. KASPRZAK

Staff: Ewa Szczęsna (PhD student)

Research profile:

Molecular motors are systems of one or several molecules, which are capable of cyclically converting chemical energy derived from adenosine triphosphate (ATP) hydrolysis into mechanical work. The motor protein Ncd, member of kinesin-14 subfamily causes sliding and expansion of an anti-parallel microtubule array by moving one microtubule over another. By hydrolyzing one ATP molecule, Ncd generates a force of up to 7 pN and makes an 8-nm step towards the minus end of the microtubule. In the cell, Ncd plays a pivotal role in the maintenance and organization of the mitotic spindle. Our group is concerned with the force generation mechanism by this motor and particularly with the role of subunit interactions in this process. Our work is also focused on the relationship between the type of motility that Ncd exhibits and its behavior and regulation in the cell. To achieve these goals we combine protein engi- neering with advanced microscopic techniques and classical biochemistry

Methods:

• protein engineering
• in vitro motility measurements of motor proteins
• total internal reflection fluorescence microscopy (TIRF)
• fluorescence resonance energy transfer (FRET)

Current research activities:

• is the ability of the mitotic kinesin Ncd to recognize the relative orientation of transported microtubules associated with the subunit structure of the motor?
• reconstruction of the microtubule plus-end tracking system of Ncd by EB1. Observation of the transport in vitro using a TIRF microscope
• mapping of the interacting regions on the Ncd-EB1 complex using point mutations in the binding areas
• identification of an Ncd domain responsible for localisation of the motor in the spindle at various stages of cell division

Selected publications:

Fink G., Hajdo L., Reuther C., Skowronek K.J., Kasprzak A.A., Diez S. (2009) The mitotic kinesin-14 Ncd, drives directional microtubule-microtubule sliding. Nature Cell Biology, 11: 717-723.

Kocik E., Skowronek K.J., Kasprzak A.A. (2009) Interactions between subunits in heterodimeric Ncd molecules. Journal of Biological Chemistry, 284: 35735-35745.

Nieznańska H., Dudek E., Zajkowski T., Szczęsna E., Kasprzak A.A., Nieznański K. (2012) Prion protein impairs kinesin-driven transport. Biochemical and Biophysical Research Communications, 425: 788-793.

Robaszkiewicz, K. Dudek E., Kasprzak A.A., Moraczewska, J. (2012) Human congenital myopathy-related mutations in TPM3 have heterogeneous functional effects. Biochimica et Biophysica Acta, 1822: 1562-1569.

Szczęsna E., Kasprzak A.A. (2012) The C-terminusof kinesin-14 Ncd is a crucial component of the force generating mechanism. FEBS Letters, 586: 854-858