Acetylation of histone H3 on lysine 56 occurs during mitotic and meiotic S phase in fungal species. This acetylation blocks a direct electrostatic interaction between histone H3 and nucleosomal DNA, and the absence of this modification is associated with extreme sensitivity to genotoxic agents. We show here that H3-K56 acetylation is catalyzed when Rtt109, a protein that lacks significant homology to known acetyltransferases, forms an active complex with either of two histone binding proteins, Asf1 or Vps75. Rtt109 binds to both these cofactors, but not to histones alone, forming enzyme complexes with kinetic parameters similar to those of known histone acetyltransferase (HAT) enzymes. Therefore, H3-K56 acetylation is catalyzed by a previously unknown mechanism that requires a complex of two proteins: Rtt109 and a histone chaperone. Additionally, these complexes are functionally distinct, with the Rtt109/Asf1 complex, but not the Rtt109/Vps75 complex, being critical for resistance to genotoxic agents.

Full structural characterization of complex mixtures such as humic acid extracts has been elusive because of insufficient compound resolution with conventional techniques. Using electrospray ionization coupled with Fourier transform ion cyclotron resonance mass spectrometry, we were able to resolve individual compounds within humic and fulvic acid mixtures (mass resolving power approximately 80000 at 300 m/z). We examined two samples in detail: (1) dissolved organic matter (primarily fulvic acids) from Suwannee River, GA, and (2) a humic acid extract from a degraded wood collected on Mt. Rainier, WA. Sample conditions (such as solvent, pH, and concentration) and instrument parameters (such as source voltages, trapping potentials, and excitation parameters) were optimized to yield the highest mass resolving power with the least mass discrimination in positive ion mode. High resolving power was achieved with low ion densities combined with coadding numerous scans. The increased resolution allowed molecular-level comparisons of the two samples which in turn could be used to estimate the relative similarity of individual compound distribution as well as an indication of the dominant diagenetic processes in the two source environments.

The extent and pattern of histone post-translational modifications is a key determinant dictating the structure of chromatin. We employed mass spectrometry to map the post-translational modifications present on mammalian core histones. Using accurate peptide mass fingerprinting on proteolytic digests of purified histones, we identified more than 20 novel sites of histone modification. One newly identified site of methylation, histone H4 lysine 59, maps to the surface of the nucleosome in close proximity to the site of the only identified histone core modification, histone H3 lysine 79. Consistent with the role of histone H3 lysine 79 methylation in the formation of silent chromatin structure, histone H4 lysine 59 is essential for transcriptional silencing at the yeast silent mating loci and telomeres.

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