Figure 6: Effect of A-lattice seam enrichment on GDP MTs.

Pure S. pombe single isoform tubulin MTs were assembled with GTP for B-lattice single-seam MTs (a,b) or with GTP and Mal3FL for A-lattice-enriched MTs (c). MTs were attached to a flow cell surface coated with rKin430, a double-headed construct of rat kinesin-1. The flow cells were flushed with buffer to remove unbound MTs and Mal3, then the MTs were imaged by dark-field microscopy, kymographs created and the shrinkage rates measured (Table 3). The B-lattice single-seam GDP MT was stable when rigor bound to rKin430 kinesin (a). Only when the B-lattice single-seam MT flow cell was flushed with buffer containing ATP to enable MT translocation was significant shrinkage observed (b). The rigor-bound A-lattice-enriched MTs were unstable and rapidly disassembled before addition of ATP (c). Blue: rigor-bound kinesin heads. Green: detached kinesin heads.