Faculty of Biological Sciences

Molecular Contractility Research

Research Interests:

Our interests range from the structure and function of the contractile apparatus of muscle; the myofibril, to the structure and functions of the plethora of myosins discovered over the last 10 years, in non-muscle cells and dynein. We use a wide range of techniques such as: wet biochemistry (protein isolation, expression and characterisation), biophysical methods (eg analytical ultracentrifugation, AFM), light microscopy (confocal, time-lapse fluorescence and deconvolution) electron microscopy, including time-resolved cryo-EM, molecular genetic tools and cell biology (tissue culture, live cell imaging) to investigate the structure and function of the muscle proteins myosin, titin and actin.We also investigate cell behaviour using time-lapse phase microscopy, and time-lapse fluorescence microscopy and Total Internal Reflection Fluorescence Microscopy (TIRFM) to image single molecules of GFP-myosin (with Dr Justin Molloy at Nat Inst for Med Res).  Our research forms part of the major effort in structural biology at Leeds.

NEW! Alternative Muscle Club Meeting at Leeds in November! More details here:

Recent Highlights:

Molecular Motors: Structure and Function.

click here for myosin v results

Myosin 5

Myosin 5, is a motor found in non-muscle cells that is involved in moving proteins and vesicles (cargo) around in cells along tracks of actin. Myosin 5 is a dimer, so it has two motor domains, and these take alternate steps along actin, to carry its cargo to the right place in the cell.

Using electron microscopy and single particle image processing we have elucidated the structure of myosin 5 attached to actin (Nature 405, 804 - 807 (15 Jun 2000)).

This movie has been made from a series of electron micrographs of myosin 5 bound to actin, so you can see how myosin 5 might 'walk' along actin.

More recently, we have investigated how this motor folds up when it is 'switched off'  and how the part of the tail that is important for folding and switching off this myosin (Nature 442, 212 - 215 (13 Jul 2006))

Myosin 6

Myosin 6 is important in making sure your hair cells (in the cochlea of the ear) work properly, so that you can hear sounds. We've recently shown that myosin 6 is a monomeric with a large working stroke (The EMBO Journal (2004) 23, 1729–1738).

The picture shows molecules of myosin 6 in the absence (B) and presence (C) of ATP. (Scale Bar, 10nm)

Myosin 7

Myosin 7

Myosin 7a is important for hearing too. We have imaged single molecules of myosin 7a from Drosophila melanogaster, and found that it is switched on and off by folding (Yang et al., PNAS (2009) 106 4189-4194)

In the picture: red is the motor domain, yellow the lever, green the tail except for the C-terminal FERM domain which is in blue. The tip of the tail (the last 100 amino acids, or 1/3rd of the FERM domain) is required to regulate Myosin 7a activity.

 


The image shows a model of myosin 10 and the SAH domain, and how it might lengthen the lever, and help this myosin take big steps when it walks on actin.

Myosin 10

Many myosins are predicted to be monomers or dimers, based on whether or not the amino acid sequence contains a region predicted to form a coiled coil. If a region of coiled coil is present, then this can dimerise myosins.  However, coiled coil prediction programmes don't work very well for highly charged regions, and it turns out that some myosins (myosin 6, 7a, 10, and myoM (Dictyostelium) have a highly charged region just after the neck/lever of the motor, where you would normally find coiled coils in other myosins.  We've now shown that this region in myosin 10 does not form coiled coil but is actually a stable single alpha helix (or SAH) domain (J. Biol. Chem. 2005, vol. 280, no41, pp. 34702-3470) and that this myosin is monomeric.

Find out more here.

Cover of Cell

Dynein

Dynein, is a motor found in many different types of cell that moves cargo along tracks made from microtubules.  It is a large protein that is difficult to crystalise, but our group recently used electron microscopy and single particle image processing to determine the structure and power stroke of dynein (Nature 421, 715-718 (13 February 2003)).

and see 'AAA+ Ring and Linker Swing Mechanism in the Dynein Motor ' (2009) by Roberts et al., Cell, 136, 485-496

Building a Muscle Sarcomere - a 'titin-ic' effort.

When muscle forms, sarcomeres - the repeating contractile units of muscle - have to be built from scratch. We've just shown that part of a giant protein called titin is essential for building the muscle sarcomeres. (J Cell Sci 2006;119 4322-4331)

The picture shows a mouse embryonic stem cell that is differentiated into a beating cardiomyocyte, and then stained and imaged using a deconvolution microscope to show the regular stripy appearance of normal muscle sarcomeres.

Find out more here.

More Movies....

This is an animation of an atomic model of the head of the myosin 2 molecule docked onto actin in two conformations thought to mimic the start (shown) and end of the power stroke. (Shockwave plugin available here).

Or you can download the movie (6.0MB)

 

 

Myofibrils from rabbit skeletal muscle contracting when irrigated with ATP and calcium ions.

This is the classical experiment of Huxley and Hanson (Nature (1954) 173, 973),  that formed a cornerstone of the sliding filament mechanism of contraction. Unconstrained by the skeleton, the myofibrils contract irreversibly to about one tenth their starting length, dramatically showing the power of the contractile machinery. They are seen here (as they were by Huxley and Hanson) using phase contrast light microscopy: a method which allows structure to be seen without any damaging staining. In the Leeds Molecular Contractility group we are investigating how the remarkably regular myofibril structure is created and maintained, and how the contractile machinery works at the atomic level.


Pre- and post-power stroke conformations of dynein.


For more see http://www.fbs.leeds.ac.uk/research/contractility/dynein/index.htm

 

Find out more about our research by going to our individual webpages!

(Professor John Trinick, Dr Peter Knight, Dr Michelle Peckham and Dr Stan Burgess)

111Mb, Quicktime 6Mb, Quicktime


Vacancies

We are actively recruiting post-docs and graduate students to join our group to contribute to a range of projects on myosin, actin and titin. The Leeds Muscle Group is lively and friendly and interested in application of state-of-the-art methods to important questions in muscle and motility.  A background in biological sciences is not essential and we are also keen to recruit from other disciplines such as physics or mathematics.


 

The Dynein homepage is now at www.dynein.org

 

 

111Mb, Quicktime 6Mb, Quicktime