A 22 kDa protein has been isolated from maize seeds that has potent activity against a number of clinically important human pathogens. In addition, this protein, named zeamatin, has the unique ability to synergize with known antifungal drugs, decreasing the dose required for fungal-cell killing while increasing the spectrum of activity. We propose, as a prelude to in vivo testing, to determine the toxicity of zeamatin and zeamatin-antifungal drug combinations on mammalian skin, cervix and kidney tissue-culture cells. We will also determine the efficacy of zeamatin and zeamatin-drug combinations in killing fungi in tissue-culture. Finally, we will determine if peptide fragments of zeamatin have antifungal activity. GRANT =P01HG00205 The goal of the Center is to determine the nucleotide sequence of the entire genome (ca. 12.5 million basepairs of DNA) in Saccharomyces cerevisae in five years and to relate as much of it as possible to simple measures of biological function, The sequence itself will be begun using novel and efficient strategies for large-scale sequencing from existing, ordered 15 kb (average insert) lambda phage clones, yet using basically current (1990) technology. The first-generation strategy is based on low-multiplicity multiplex sequencing followed by a newly-devised closure strategy, that orients and orders the contigs and bridges the gaps in sequence in a single step using the polymerase chain reaction (PCR). A new variation on multiplex sequencing using random transposon insertions, will be implemented. Automated sequencing methods will be applied to the bridging PCR fragments. Conservative estimates suggest that this strategy, when fully implemented, should allow a single worker to produce as much as 300 kb of useful sequence per year. A second project aims to fully correlate the genetic and physical maps of Saccharomyces with the sequence and to learn, by standard and novel molecular methods, as much as possible about the encoded proteins and their biological functions. A third project is the construction of fully integrated databases for the genome of Saccharomyces and provision of an interim mapping and sequencing service to investigators who provide convincing evidence that they have cloned a new gene from this yeast. A variety of techonological improvements in sequencing, gene-protein- function correlation, and automation are proposed as development projects. We propose a new method of organization whereby promising young scientists can work together in an academic environment on this major cooperative effort while enhancing their own abilities and careers as independent scientists. If successful, all these innovations should have a major impact on the future of large-scale endeavors in biology like the Human Genome Project.
