Prof Peter Knight

Peter Knight is Professor of Molecular Contractility in the Institute of Molecular and Cellular Biology. He studied at King's College, London University (BSc, Biology, 1971), where he was infected with an enthusiasm for the structural basis of muscle contraction by Professor Jean Hanson. He stayed on to gain a PhD (Biophysics, 1976), working with Dr Gerald Offer on structural and enzymatic properties of cross-linked actin. Postdoctoral studies on thick filament structure were with Dr Bill Harrington at the Johns Hopkins University (Baltimore, USA) and with Gerald Offer, now in Bristol. He was at Bristol from 1980, mainly as a staff scientist at the BBSRC Food Research Institute, where he applied a knowledge of muscle ultrastructure and properties to understanding the properties of muscle as a food. He joined the University of Leeds in 1997, teaches gross anatomy and muscle ultrastructure, and is a member of the Muscle Research Group.

Research Areas: Structural Basis of Movement in Cells and Tissues

Research Interests: I am fascinated by the structural basis of movement in cells and tissues. Therefore am interested in the structures of myosin and actin and the filaments they form, not only from muscle, but also including the diverse members of the myosin superfamily. How the structures change cyclically to produce movement, and how movement is controlled are especial focuses. Muscle myofibrils are assembled and maintained with great precision, and I am interested in the underlying mechanisms, such as the molecular ruler properties of titin and nebulin, and the mechanisms such as titin elasticity that maintain order. Techniques I have used to pursue these interests include electron microscopy of single molecules and macromolecular assemblies, X-ray diffraction of muscle, immunological techniques and protein biochemistry. Collaborations within and beyond the School have involved image processing, computer modelling and protein dynamics simulation.

Research projects:

Myosins are a diverse family of proteins They have in common a motor domain that interacts with filaments of F-actin, and splits ATP to power movement and force generation. The rest of the molecule of each species of myosin is adapted to its particular role in the cell. In muscle for instance, contraction is driven by myosin II, which has two motors each attached by a short neck to a long tail. The polymerisation of the tails to form bipolar filaments allows the heads to make brief, asynchronous interactions with actin without the filament as a whole detaching. This allows both fast movement when loads are light, and greater force (and slower speed) when loads are heavy. A major research interest is to understand the mechanism of this motor, the adaptations of the molecule for specific functions and the ways that its activity is regulated. Although atomic structures of the motor have been obtained by crystallography, the F-actin-myosin complex is not been amenable to this method, and there is much that electron microscopy can contribute.

The progress that can come from microscopy is exemplified by a recent collaboration with Prof John Trinick and other members of the Leeds Muscle Group, together with Dr Jim Sellers and colleagues at NIH (USA). We used electron microscopy and image analysis to show that myosin V, which has two unusually long necks attached to a tail, can take sufficiently long steps along F-actin that the underlying helical arrangement of actin subunits can be ignored and the molecule can walk in a straight line. A function of this myosin is probably to carry vesicle cargoes attached to the end of its tail, so the linear walk may be an adaptation that avoids the cargo colliding with other cellular structures.

The motor structure is also tuned so that it stays attached to actin for a much greater fraction of the ATPase cycle time, and this enables it to walk, rather than skip along the actin filament. The key feature of walking in this context is that the molecule is never detached from actin, which would be especially important if vesicles are transported by single myosin molecules. This adaptation has allowed us to get images of the myosin with both its heads attached to actin during ATP-fuelled walking. When the molecule is attached by both its heads, we find that the leading one is in a previously conjectured state at the start of its working stroke that has not been seen before. The structure of this head is consistent with the idea that myosin moves along actin by changing its shape after attaching. We are applying high resolution methods to reveal more details of this new structure and look forward to exploiting the diversity of the myosin family to understand the structural basis of myosin function. For more detail and movies of this myosin attached to actin, see the Group?s web pages at http://www.leeds.ac.uk/bms/research/muscle/muscle.htm.

To understand myosin structure better, I have collaborated with Dr Gerald Offer (Bristol) to build atomic models of myosin II constructed from the atomic structures of the isolated heads, and a modelled tail. This has shown routes for interactions between the heads that may be important for regulation of activity. It has also pointed up a paradox between the expected structure of the molecule and its known properties. The model shows the heads closely associated at their junction with the tail and therefore sterically constrained, whereas the heads are known to be highly mobile on a microsecond time scale. The solution of this may be that the start of the tail, which is a two chain a-helical coiled coil, may be a less stable structure than widely thought. Unfolding of some a-helix may allow it to function, for instance, as the series elastic component that is required in each molecule to allow it to function independently within the thick filament structure. Evidence for this dynamic role of the tail has come from my observation by electron microscopy of separation of the heads and shortening of the tail in molecules reacted with an antibody directed against the first part of the tail.

We have used this myosin model in collaboration with Drs Raul Padron and Lorenzo Alamo (Caracas, Venezuela) to interpret 3D reconstructions of myosin filaments imaged in the electron microscope. We used the thick filaments from Tarantula muscle as test objects, because when they are switched off (as in relaxed muscle), the heads are arranged into a particularly well-ordered lattice. The structure of the thick filament has eluded detailed description, but our model is able to fit this surface lattice of heads well. This indicates that the model structure may be favoured when activity is inhibited. The arrangement of the heads on the filament suggests sites of interaction between neighbouring molecules in the filament that may have a role in regulating activity.

Many cells contain more than one member of the myosin family. Where and how these are distributed and controlled, and the functions they perform are all still largely unknown, and I am collaborating with Dr Michelle Peckham in the Muscle Group to explore these topics by a combination of fluorescence and electron microscopy.

See http://www.leeds.ac.uk/bms/research/muscle/peckham/indexnew.html

Titin is a giant, multifunctional, muscle protein responsible for passive muscle tension and myofibrillar assembly. I am interested particularly in its function as a ruler for dictating the assembly of myosin molecules and other proteins to make the thick filaments. The amino acid sequence and modular structure of titin suggest that it contains binding sites for several thick filament proteins including myosin and C-protein, and in this way ensures that their assembly produces the remarkably exact structure that is found. Direct evidence for this attractive hypothesis remains scant however, and so we are testing it using constructs of titin and fragments of myosin.