ABSTRACT
In eukaryotes, several lysine residues on histone proteins are methylated. This posttranslational modification is linked to a myriad of nuclear-based transactions such as epigenetic inheritance of heterochromatin, regulation of gene expression, DNA damage repair, and DNA replication. The majority of the enzymes responsible for writing these marks onto chromatin belong to the SET domain family of histone lysine methyltransferases. Although they often share important structural features, including a conserved catalytic domain, SET domain enzymes use different mechanisms to achieve substrate recognition, mono-, di-, or trimethylate lysine residues and some require other proteins to achieve maximal methyltransferase activity. In this chapter, we summarize our efforts to purify, crystallize, and enzymatically characterize SET domain enzymes with a specific focus on the histone H3K27 monomethyltransferase ATXR5.
Subject(s)
Histone-Lysine N-Methyltransferase/chemistry , Histone-Lysine N-Methyltransferase/metabolism , Amino Acid Sequence , Animals , Crystallography, X-Ray/methods , Enzyme Assays/methods , Escherichia coli/genetics , Histone-Lysine N-Methyltransferase/genetics , Histone-Lysine N-Methyltransferase/isolation & purification , Humans , Models, Molecular , Plant Proteins/chemistry , Plant Proteins/genetics , Plant Proteins/isolation & purification , Plant Proteins/metabolism , Protein Domains , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Ricinus/chemistry , Ricinus/genetics , Ricinus/metabolism , Sequence Alignment , Up-RegulationABSTRACT
The influence of processing parameters and synthetic strategies in the properties of sol-gel derived silica matrices intended for the release of bioactive compounds was investigated. The time-evolution of the matrix properties during its aging at room temperature in the dry and wet forms was investigated by measuring some of its physical and drug retaining properties. The results indicate that long term gel aging in the wet form is fundamental for the obtainment of dry matrices that are stable upon storage, a fundamental requirement for any practical application. In the case of hybrid matrices obtained by replacing part of the tetraethoxysilane precursor with mono-methyl trimethoxysilane, the order of addition of the reaction component is also important in determining the properties of the final dry gel, probably by influencing the polymer structural properties. This parameter acts synergistically with the matrix composition in determining the release properties of xerogels embedded with bioactive compounds.