MCM2 and MCM3 are essential genes believed to play important roles in the initiation of DNA replication in Saccharomyces cerevisiae. Mutants defective in Mcm2 or Mcm3 are remarkably similar in phenotype. They both show an autonomously replicating sequence (ARS)-specific minichromosome maintenance defect, although their ARS specificities are not identical. In addition, these mutants exhibit a premitotic cell cycle arrest and an increase in chromosome loss and recombination. Genetic studies suggest that the two MCM gene products play interacting or complementary roles in DNA replication. Double mutants of mcm2-1 and mcm3-1 are inviable at the permissive growth temperature (23 degrees C) for each of the single mutants. Furthermore, overproduction of Mcm3 accentuates the deleterious effect of the mcm2-1 mutation, whereas overproduction of Mcm2 partially complements the mcm3-1 mutation. MCM2 encodes a protein of 890 amino acids containing a putative zinc-finger domain that is essential for Mcm2 function. Mcm2 shows striking homology to Mcm3 and three other proteins, Cdc46 of S. cerevisiae, and Nda4 and Cdc21 of Schizosaccharomyces pombe. The phenotypes of mutants defective in these proteins suggest that they belong to a protein family involved in the early steps of DNA replication.