One-Atom Substitution Enables Direct and Continuous Monitoring of Histone Deacylase Activity

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F86652036%3A_____%2F19%3A00519596" target="_blank" >RIV/86652036:_____/19:00519596 - isvavai.cz</a>

Alternative codes found

RIV/00216224:14310/19:00113392

Result on the web

<a href="https://pubs.acs.org/doi/10.1021/acs.biochem.9b00786" target="_blank" >https://pubs.acs.org/doi/10.1021/acs.biochem.9b00786</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1021/acs.biochem.9b00786" target="_blank" >10.1021/acs.biochem.9b00786</a>

Result language

angličtina

Original language name

One-Atom Substitution Enables Direct and Continuous Monitoring of Histone Deacylase Activity

Original language description

We developed a one-step direct assay for the determination of histone deacylase (HDAC) activity by substituting the carbonyl oxygen of the acyl moiety with sulfur, resulting in thioacylated lysine side chains. This modification is recognized by class I HDACs with different efficiencies ranging from not accepted for HDAC1 to kinetic constants similar to that of the parent oxo substrate for HDAC8. Class II HDACs can hydrolyze thioacylated substrates with approximately 5-10-fold reduced k(cat) values, which resembles the effect of thioamide substitution in metallo-protease substrates. Class IV HDAC11 accepts thiomyristoyl modification less efficiently with an similar to 5-fold reduced specificity constant. On the basis of the unique spectroscopic properties of thioamide bonds (strong absorption in spectral range of 260-280 nm and efficient fluorescence quenching), HDAC-mediated cleavage of thioamides could be followed by ultraviolet-visible and fluorescence spectroscopy in a continuous manner. The HDAC activity assay is compatible with microtiter plate-based screening formats up to 1536-well plates with Z' factors of >0.75 and signal-to-noise ratios of >50. Using thioacylated lysine residues in p53-derived peptides, we optimized substrates for HDAC8 with a catalytic efficiency of >250000 M-1 s(-1), which are more than 100-fold more effective than most of the known su bstrates. We determined inhibition constants of several inhibitors for human HDACs using thioacylated peptidic substrates and found good correlation with the values from the literature. On the other hand, we could introduce N-methylated, N-acylated lysine residues as inhibitors for HDACs with an IC50 value of 1 mu M for an N-methylated, N-myristoylated peptide derivative and human HDAC11.

Czech name

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Czech description

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Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

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OECD FORD branch

10608 - Biochemistry and molecular biology

Project

Result was created during the realization of more than one project. More information in the Projects tab.

Continuities

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Publication year

2019

Confidentiality

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Name of the periodical

Biochemistry

ISSN

0006-2960

e-ISSN

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Volume of the periodical

58

Issue of the periodical within the volume

48

Country of publishing house

US - UNITED STATES

Number of pages

13

Pages from-to

4777-4789

UT code for WoS article

000500838300002

EID of the result in the Scopus database

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