Alanine scanning of Arp1 delineates a putative binding site for Jnm1/dynamitin and Nip100/p150Glued.
Arp1p is the only actin-related protein (ARP) known to form actin-like filaments. Unlike actin, Arp1p functions with microtubules, as part of the dynein regulator, dynactin. Arp1p's dissimilar functions imply interactions with a distinct set of proteins. To distinguish surface features relating to Arp1p's core functions and to identify the footprint of protein interactions essential for dynactin function, we performed the first complete charge-cluster-to-alanine scanning mutagenesis of an ARP and compared the results with a similar study of actin. The Arp1p mutations revealed three nonoverlapping surfaces with distinct genetic properties. One of these surfaces encompassed a region unique to Arp1p that is crucial for Jnm1p (dynamitin/p50) and Nip100p (p150(Glued)) association as well as pointed-end associations. Unlike the actin mutations, none of the ARP1 alleles disrupt filament formation; however, one pointed-end allele delayed the elution of Arp1p on gel filtration, consistent with loss of additional subunits.
Clark
Sean W
SW
Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA.
Rose
Mark D
MD
D000199:Actins / Q000235:genetics / Q000378:metabolism; D000409:Alanine / Q000235:genetics* / Q000378:metabolism; D000483:Alleles; D001483:Base Sequence; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D000072159:Dynactin Complex; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D009711:Nucleotides / Q000378:metabolism; D010641:Phenotype; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
C089021:ARP1 protein, S cerevisiae; C465210:Act1 protein, S cerevisiae; D000199:Actins; D003598:Cytoskeletal Proteins; D000072159:Dynactin Complex; C086450:JNM1 protein, S cerevisiae; D008869:Microtubule-Associated Proteins; C000606366:NIP100 protein, S cerevisiae; D009711:Nucleotides; D029701:Saccharomyces cerevisiae Proteins; D000409:Alanine