title: Structural changes in Mcm5 protein bypass Cdc7-Dbf4 function and reduce replication origin efficiency in Saccharomyces cerevisiae.
abstract: Eukaryotic chromosomal replication is a complicated process with many origins firing at different efficiencies and times during S phase. Prereplication complexes are assembled on all origins in G(1) phase, and yet only a subset of complexes is activated during S phase by DDK (for Dbf4-dependent kinase) (Cdc7-Dbf4). The yeast mcm5-bob1 (P83L) mutation bypasses DDK but results in reduced intrinsic firing efficiency at 11 endogenous origins and at origins located on minichromosomes. Origin efficiency may result from Mcm5 protein assuming an altered conformation, as predicted from the atomic structure of an archaeal MCM (for minichromosome maintenance) homologue. Similarly, an intragenic mutation in a residue predicted to interact with P83L suppresses the mcm5-bob1 bypass phenotype. We propose DDK phosphorylation of the MCM complex normally results in a single, highly active conformation of Mcm5, whereas the mcm5-bob1 mutation produces a number of conformations, only one of which is permissive for origin activation. Random adoption of these alternate states by the mcm5-bob1 protein can explain both how origin firing occurs independently of DDK and why origin efficiency is reduced. Because similar mutations in mcm2 and mcm4 cannot bypass DDK, Mcm5 protein may be a unique Mcm protein that is the final target of DDK regulation.
authors.affiliation: Department of Genome Sciences, University of Washington, Seattle, USA.
authors.forename: Margaret L
authors.initials: ML
authors.lastname: Hoang
authors.forename: Ronald P
authors.initials: RP
authors.lastname: Leon
authors.forename: Luis
authors.initials: L
authors.lastname: Pessoa-Brandao
authors.forename: Sonia
authors.initials: S
authors.lastname: Hunt
authors.forename: M K
authors.initials: MK
authors.lastname: Raghuraman
authors.forename: Walton L
authors.initials: WL
authors.lastname: Fangman
authors.forename: Bonita J
authors.initials: BJ
authors.lastname: Brewer
authors.forename: Robert A
authors.initials: RA
authors.lastname: Sclafani
mesh_terms: D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone; D015825:Chromosomes, Fungal / Q000378:metabolism; D004261:DNA Replication; D004271:DNA, Fungal / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D005656:Fungal Proteins / Q000235:genetics; D064148:Minichromosome Maintenance Complex Component 4; D009154:Mutation / Q000235:genetics; D010957:Plasmids; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D018741:Replication Origin; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; C088096:Dbf4 protein, S cerevisiae; D005656:Fungal Proteins; C069200:MCM5 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C489404:CDC7 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C069199:MCM2 protein, S cerevisiae; C092802:MCM4 protein, S cerevisiae; D064148:Minichromosome Maintenance Complex Component 4