Unusual properties of the fungal conventional kinesin neck domain from Neurospora crassa.
Fungal conventional kinesins are unusually fast microtubule motor proteins. To compare the functional organization of fungal and animal conventional kinesins, a set of C-terminal deletion mutants of the Neurospora crassa conventional kinesin, NcKin, was investigated for its biochemical and biophysical properties. While the shortest, monomeric construct comprising the catalytic core and the neck-linker (NcKin343) displays very high steady-state ATPase (k(cat) = 260/s), constructs including both the full neck and adjacent hinge domains (NcKin400, NcKin433 and NcKin480) show wild-type behaviour: they are dimeric, show fast gliding and slower ATP turnover rates (k(cat) = 60-84/s), and are chemically processive. Unexpectedly, a construct (NcKin378, corresponding to Drosophila KHC381) that includes just the entire coiled-coil neck is a monomer. Its ATPase activity is slow (k(cat) = 27/s), and chemical processivity is abolished. Together with a structural analysis of synthetic neck peptides, our data demonstrate that the NcKin neck domain behaves differently from that of animal conventional kinesins and may be tuned to drive fast, processive motility.
Kallipolitou
A
A
Adolf-Butenandt-Institute, Cell Biology 1b, Universität München, Schillerstrasse 42, D-80336 Munich, Germany.
Deluca
D
D
Majdic
U
U
Lakämper
S
S
Cross
R
R
Meyhöfer
E
E
Moroder
L
L
Schliwa
M
M
Woehlke
G
G
D000244:Adenosine Diphosphate / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D020134:Catalytic Domain; D002465:Cell Movement; D002942:Circular Dichroism; D003001:Cloning, Molecular; D003545:Cysteine / Q000737:chemistry; D004330:Drosophila; D016547:Kinesins / Q000737:chemistry*; D007700:Kinetics; D013058:Mass Spectrometry; D008870:Microtubules / Q000378:metabolism; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D009492:Neurospora crassa / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013696:Temperature; D013997:Time Factors
D010455:Peptides; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D016547:Kinesins; D003545:Cysteine