It has been known since the 1940s that such products can include intramolecular and intermolecular crosslinked species and that the reaction conditions ( e.g. Initially using amino acids, and subsequently proteins and other substrates, it was shown that formaldehyde reacts in vitro with a wide range of functional groups, forming a complex array of products ( 6, 7). The chemical complexity of formaldehyde-mediated reaction products was appreciated 70 years ago ( 5). Progress in capturing crosslinked complexes will also be discussed with an emphasis on the impact of a better understanding of formaldehyde chemistry in vivo.įormaldehyde is the smallest aldehyde, an electrophilic molecule susceptible to chemical attack by a wide range of nucleophilic species of biological interest. Beginning with basic chemical reactivity, this review will explore the ability of formaldehyde to crosslink with proteins and DNA to form protein-protein or protein-DNA complexes, common molecular quenchers, and the potential for crosslink reversal. The following discussion provides a framework for understanding aspects of formaldehyde function when used to trap macromolecular complexes in cells, with the main features shown in Fig. Development of experimental strategies to achieve these goals will require a deeper and more comprehensive understanding of the effects mediated by formaldehyde in cells. A major goal of ongoing work is to understand kinetic and thermodynamic aspects of chromatin complex assembly at single copy loci in vivo. The analysis of formaldehyde-fixed chromatin has provided fundamental insights into where and when regulatory factors associate with the DNA template in vivo, but it in general does not provide unambiguous information about chromatin binding kinetics. These issues are of significance for designing crosslinking-based studies as well as for properly interpreting the resulting data. This information provides a basis for understanding how formaldehyde functions in widely used assays in the chromatin field, and conversely, highlights less well understood aspects of formaldehyde behavior in cells. Here, we briefly review prior work describing formaldehyde reactivity toward proteins, DNA, and their constituent monomers. A recent complementary perspective highlights gaps in knowledge with a particular focus on how formaldehyde crosslinking data have been used to interpret aspects of chromatin three-dimensional organization ( 4). In this review, we focus on its use in chromatin immunoprecipitation approaches and protein-protein interaction studies applied to understand the location and abundance of transcription factor binding along DNA. Prior to its use in the chromatin field, formaldehyde use had a long history in a number of fields, including vaccine production ( 1, 2) and histology ( 3).
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