The yeast histone acetyltransferase A2 complex, but not free Gcn5p, binds stably to nucleosomal arrays.
We have investigated the structural basis for the differential catalytic function of the yeast Gcn5p-containing histone acetyltransferase (HAT) A2 complex and free recombinant yeast Gcn5p (rGcn5p). HAT A2 is shown to be a unique complex that contains Gcn5p, Ada2p, and Ada3p, but not proteins specific to other related HAT A complexes, e.g. ADA, SAGA. Nevertheless, HAT A2 produces the same unique polyacetylation pattern of nucleosomal substrates reported previously for ADA and SAGA, demonstrating that proteins specific to the ADA and SAGA complexes do not influence the enzymatic activity of Gcn5p within the HAT A2 complex. To investigate the role of substrate interactions in the differential behavior of free and complexed Gcn5p, sucrose density gradient centrifugation was used to characterize the binding of HAT A2 and free rGcn5p to intact and trypsinized nucleosomal arrays, H3/H4 tetramer arrays, and nucleosome core particles. We find that HAT A2 forms stable complexes with all nucleosomal substrates tested. In distinct contrast, rGcn5p does not interact stably with nucleosomal arrays, despite being able to specifically monoacetylate the H3 N terminus of nucleosomal substrates. Our data suggest that the ability of the HAT A2 complex to bind stably to nucleosomal arrays is functionally related to both local and global acetylation by the complexed and free forms of Gcn5p.
Sendra
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R
Departament de Bioquimica i Biologia Molecular, Universitat de València, E-46100 València, Spain.
Tse
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C
Hansen
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JC
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000107:Acetylation; D000123:Acetyltransferases / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002499:Centrifugation, Density Gradient; D002645:Chickens; D002843:Chromatin / Q000378:metabolism*; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000302:isolation & purification / Q000378:metabolism*; D051548:Histone Acetyltransferases; D006657:Histones / Q000097:blood / Q000378:metabolism; D009707:Nucleosomes / Q000378:metabolism*; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity
D002843:Chromatin; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D006657:Histones; D009707:Nucleosomes; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D000105:Acetyl Coenzyme A; D000123:Acetyltransferases; D051548:Histone Acetyltransferases; D011494:Protein Kinases