Lysine deacetylases, like histone deacetylases (HDACs) and sirtuins (SIRTs), are involved in many regulatory processes such as control of metabolic pathways, DNA repair, and stress responses. Besides robust deacetylase activity, sirtuin isoforms SIRT2 and SIRT3 also show demyristoylase activity. Interestingly, most of the inhibitors described so far for SIRT2 are not active if myristoylated substrates are used. Activity assays with myristoylated substrates are either complex because of coupling to enzymatic reactions or time-consuming because of discontinuous assay formats. Here we describe sirtuin substrates enabling direct recording of fluorescence changes in a continuous format. Fluorescence of the fatty acylated substrate is different when compared to the deacylated peptide product. Additionally, the dynamic range of the assay could be improved by the addition of bovine serum albumin, which binds the fatty acylated substrate and quenches its fluorescence. The main advantage of the developed activity assay is the native myristoyl residue at the lysine side chain avoiding artifacts resulting from the modified fatty acyl residues used so far for direct fluorescence-based assays. Due to the extraordinary kinetic constants of the new substrates (KM values in the low nM range, specificity constants between 175,000 and 697,000 M-1s-1) it was possible to reliably determine the IC50 and Ki values for different inhibitors in the presence of only 50 pM of SIRT2 using different microtiter plate formats.

• Klíčová slova
• bovine serum albumin effect, continuous activity assay, fluorescence quenching, histone deacetylases, myristoylated substrates, sirtuin inhibitors, sirtuins,
• MeSH
• barvicí látky MeSH
• lysin MeSH
• peptidy MeSH
• sirtuin 1 metabolismus   MeSH
• sirtuin 2 metabolismus   MeSH
• sirtuin 3 * metabolismus   MeSH
• sirtuiny * metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• barvicí látky MeSH
• lysin MeSH
• peptidy MeSH
• sirtuin 1 MeSH
• sirtuin 2 MeSH
• sirtuin 3 * MeSH
• sirtuiny * MeSH

This review summarizes the current knowledge on essential vitamins B1, B2, B3, and B5. These B-complex vitamins must be taken from diet, with the exception of vitamin B3, that can also be synthetized from amino acid tryptophan. All of these vitamins are water soluble, which determines their main properties, namely: they are partly lost when food is washed or boiled since they migrate to the water; the requirement of membrane transporters for their permeation into the cells; and their safety since any excess is rapidly eliminated via the kidney. The therapeutic use of B-complex vitamins is mostly limited to hypovitaminoses or similar conditions, but, as they are generally very safe, they have also been examined in other pathological conditions. Nicotinic acid, a form of vitamin B3, is the only exception because it is a known hypolipidemic agent in gram doses. The article also sums up: (i) the current methods for detection of the vitamins of the B-complex in biological fluids; (ii) the food and other sources of these vitamins including the effect of common processing and storage methods on their content; and (iii) their physiological function.

• Klíčová slova
• essential, niacin, pantothenic acid, riboflavin, thiamine,
• MeSH
• avitaminóza * MeSH
• lidé MeSH
• thiamin MeSH
• vitamin A MeSH
• vitamin B komplex * MeSH
• vitamin K MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• thiamin MeSH
• vitamin A MeSH
• vitamin B komplex * MeSH
• vitamin K MeSH

Tumor suppressors represent a critical line of defense against tumorigenesis. Their mechanisms of action and the pathways they are involved in provide important insights into cancer progression, vulnerabilities, and treatment options. Although nuclear and cytosolic tumor suppressors have been extensively investigated, relatively little is known about tumor suppressors localized within the mitochondria. However, recent research has begun to uncover the roles of these important proteins in suppressing tumorigenesis. Here, we review this newly developing field and summarize available information on mitochondrial tumor suppressors.

• MeSH
• buněčná diferenciace genetika   MeSH
• energetický metabolismus * MeSH
• lidé MeSH
• mitochondriální geny * MeSH
• mitochondrie genetika   metabolismus   MeSH
• nádorová transformace buněk genetika   metabolismus   MeSH
• nádory genetika   metabolismus   patologie   MeSH
• progrese nemoci MeSH
• regulace genové exprese u nádorů MeSH
• tumor supresorové geny * MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Histone deacetylase 11 (HDAC11) preferentially removes fatty acid residues from lysine side chains in a peptide or protein environment. Here, we report the development and validation of a continuous fluorescence-based activity assay using an internally quenched TNFα-derived peptide derivative as a substrate. The threonine residue in the +1 position was replaced by the quencher amino acid 3'-nitro-l-tyrosine and the fatty acyl moiety substituted by 2-aminobenzoylated 11-aminoundecanoic acid. The resulting peptide substrate enables fluorescence-based direct and continuous readout of HDAC11-mediated amide bond cleavage fully compatible with high-throughput screening formats. The Z'-factor is higher than 0.85 for the 15 μM substrate concentration, and the signal-to-noise ratio exceeds 150 for 384-well plates. In the absence of NAD+, this substrate is specific for HDAC11. Reevaluation of inhibitory data using our novel assay revealed limited potency and selectivity of known HDAC inhibitors, including Elevenostat, a putative HDAC11-specific inhibitor.

• Publikační typ
• časopisecké články MeSH