Abstract：
:Enzymes in the α-d-phosphohexomutase (PHM) superfamily catalyze a multistep reaction, entailing two successive phosphoryl transfers. Key to this reaction is a conserved phosphoserine in the active site, which serves alternately as a phosphoryl donor and acceptor during the catalytic cycle. In addition to its role in the enzyme mechanism, the phosphorylation state of the catalytic phosphoserine has recently been found to have widespread effects on the structural flexibility of enzymes in this superfamily. These effects must be carefully accounted for when assessing other perturbations to these enzymes, such as mutations or ligand binding. In this chapter, we focus on methods for assessing and modulating the phosphorylation state of the catalytic serine, as well as straightforward ways to probe the impacts of this modification on protein structure/flexibility. This knowledge is essential for producing homogeneous and stable samples of these proteins for biophysical studies. The methods described herein should be widely applicable to enzymes across the PHM superfamily and may also be useful in characterizing the effects of posttranslational modifications on other proteins.
journal_name
Methods Enzymoljournal_title
Methods in enzymologyauthors
Stiers KM,Beamer LJdoi
10.1016/bs.mie.2018.04.003subject
Has Abstractpub_date
2018-01-01 00:00:00pages
241-267eissn
0076-6879issn
1557-7988pii
S0076-6879(18)30137-Xjournal_volume
607pub_type
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