skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Engineering of a novel Ca{sup 2+}-regulated kinesin molecular motor using a calmodulin dimer linker

Abstract

Highlights: Black-Right-Pointing-Pointer Engineered kinesin-M13 and calmodulin involving single cysteine were prepared. Black-Right-Pointing-Pointer CaM mutant was cross-linked to dimer by bifunctional thiol reactive reagent. Black-Right-Pointing-Pointer Kinesin-M13 was dimerized via CaM dimer in the presence of calcium. Black-Right-Pointing-Pointer Function of the engineered kinesin was regulated by a Ca{sup 2+}-calmodulin dimer linker. -- Abstract: The kinesin-microtubule system holds great promise as a molecular shuttle device within biochips. However, one current barrier is that such shuttles do not have 'on-off' control of their movement. Here we report the development of a novel molecular motor powered by an accelerator and brake system, using a kinesin monomer and a calmodulin (CaM) dimer. The kinesin monomer, K355, was fused with a CaM target peptide (M13 peptide) at the C-terminal part of the neck region (K355-M13). We also prepared CaM dimers using CaM mutants (Q3C), (R86C), or (A147C) and crosslinkers that react with cysteine residues. Following induction of K355-M13 dimerization with CaM dimers, we measured K355-M13 motility and found that it can be reversibly regulated in a Ca{sup 2+}-dependent manner. We also found that velocities of K355-M13 varied depending on the type and crosslink position of the CaM dimer used; crosslink length also had a moderate effect onmore » motility. These results suggest Ca{sup 2+}-dependent dimerization of K355-M13 could be used as a novel molecular shuttle, equipped with an accelerator and brake system, for biochip applications.« less

Authors:
 [1];  [1]
  1. Department of Bioinformatics, Faculty of Engineering, Soka University, Hachioji, Tokyo 192-8577 (Japan)
Publication Date:
OSTI Identifier:
22207921
Resource Type:
Journal Article
Journal Name:
Biochemical and Biophysical Research Communications
Additional Journal Information:
Journal Volume: 423; Journal Issue: 2; Other Information: Copyright (c) 2012 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0006-291X
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; CALCIUM IONS; CALMODULIN; COMPUTER-AIDED MANUFACTURING; CYSTEINE; DIMERIZATION; DIMERS; EGTA; ENZYME ACTIVITY; ETHANE; HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY; MICROTUBULES; PEPTIDES; RABBIT TUBES

Citation Formats

Shishido, Hideki, and Maruta, Shinsaku, E-mail: maruta@soka.ac.jp. Engineering of a novel Ca{sup 2+}-regulated kinesin molecular motor using a calmodulin dimer linker. United States: N. p., 2012. Web. doi:10.1016/J.BBRC.2012.05.135.
Shishido, Hideki, & Maruta, Shinsaku, E-mail: maruta@soka.ac.jp. Engineering of a novel Ca{sup 2+}-regulated kinesin molecular motor using a calmodulin dimer linker. United States. doi:10.1016/J.BBRC.2012.05.135.
Shishido, Hideki, and Maruta, Shinsaku, E-mail: maruta@soka.ac.jp. Fri . "Engineering of a novel Ca{sup 2+}-regulated kinesin molecular motor using a calmodulin dimer linker". United States. doi:10.1016/J.BBRC.2012.05.135.
@article{osti_22207921,
title = {Engineering of a novel Ca{sup 2+}-regulated kinesin molecular motor using a calmodulin dimer linker},
author = {Shishido, Hideki and Maruta, Shinsaku, E-mail: maruta@soka.ac.jp},
abstractNote = {Highlights: Black-Right-Pointing-Pointer Engineered kinesin-M13 and calmodulin involving single cysteine were prepared. Black-Right-Pointing-Pointer CaM mutant was cross-linked to dimer by bifunctional thiol reactive reagent. Black-Right-Pointing-Pointer Kinesin-M13 was dimerized via CaM dimer in the presence of calcium. Black-Right-Pointing-Pointer Function of the engineered kinesin was regulated by a Ca{sup 2+}-calmodulin dimer linker. -- Abstract: The kinesin-microtubule system holds great promise as a molecular shuttle device within biochips. However, one current barrier is that such shuttles do not have 'on-off' control of their movement. Here we report the development of a novel molecular motor powered by an accelerator and brake system, using a kinesin monomer and a calmodulin (CaM) dimer. The kinesin monomer, K355, was fused with a CaM target peptide (M13 peptide) at the C-terminal part of the neck region (K355-M13). We also prepared CaM dimers using CaM mutants (Q3C), (R86C), or (A147C) and crosslinkers that react with cysteine residues. Following induction of K355-M13 dimerization with CaM dimers, we measured K355-M13 motility and found that it can be reversibly regulated in a Ca{sup 2+}-dependent manner. We also found that velocities of K355-M13 varied depending on the type and crosslink position of the CaM dimer used; crosslink length also had a moderate effect on motility. These results suggest Ca{sup 2+}-dependent dimerization of K355-M13 could be used as a novel molecular shuttle, equipped with an accelerator and brake system, for biochip applications.},
doi = {10.1016/J.BBRC.2012.05.135},
journal = {Biochemical and Biophysical Research Communications},
issn = {0006-291X},
number = 2,
volume = 423,
place = {United States},
year = {2012},
month = {6}
}