Sirtuins and ADP ribosyltransferases (ARDTs) are two families of related enzymes that have important medically relevant functions. For example, SIRT3, a mitochondrial sirtuin, controls mitochondrial function, and thereby exerts a strong effect on metabolism, obesity, type 2 diabetes, and metabolic syndrome. There is a major need to have simple assays to measure the activity of SIRT3, as well as other sirtuins and their close relatives, the ARDTs. Each of these enzymes uses nicotinamide adenine dinucleotide (NAD), and generates nicotinamide as a byproduct of these reactions. Unfortunately, current methods for assaying the activity of these enzymes are not optimal because they are not homogeneous, are not continuous, lack sensitivity, and often not generalizable to screening all of the diverse enzymes that generate nicotinamide. A more straightforward assay would be to directly measure the products of these enzymes in a homogeneous format. Therefore, assays that directly detect the appearance of nicotinamide would be useful for measuring the activity these enzymes, regardless of the specific substrate. Using a patented sensor technology developed by Lucerna, we will develop a universal sirtuin/ARDT assay based on a simple, homogeneous fluorescence sensor that is highly specific for nicotinamide. This Phase I SBIR application describes proof-of-concept experiments for developing, characterizing, and optimizing our nicotinamide sensors in an assay for SIRT3 activity. For Phase II, our assay will be optimized to comply with standards required for ensuring that this assay can be packaged and used in a kit format for a variety of sirtuins and related enzymes.

Public Health Relevance

Sirtuins and ADP ribosyltransferases are important classes of enzymes that affect many medically important processes, including cancer, inflammation, diabetes, and metabolic syndrome. Here we describe the development of a kit that can be used to assay these enzymes. This will result in an important research tool that will facilitate new discoveries about these important enzymes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Small Business Innovation Research Grants (SBIR) - Phase I (R43)
Project #
1R43GM105153-01A1
Application #
8591776
Study Section
Special Emphasis Panel (ZRG1-IMST-L (11))
Program Officer
Gerratana, Barbara
Project Start
2013-09-18
Project End
2014-12-31
Budget Start
2013-09-18
Budget End
2014-08-31
Support Year
1
Fiscal Year
2013
Total Cost
$211,753
Indirect Cost
Name
Lucerna, Inc.
Department
Type
DUNS #
961919508
City
New York
State
NY
Country
United States
Zip Code
10065