Development and Validation of a Three-Step Screening Strategy for Extracellular Salt-Tolerant Nucleases From Marine...

Area of Science:

• Marine microbiology
• Enzymology
• Biotechnology

Background:

• Extracellular nucleases are crucial enzymes with diverse applications.
• Screening marine organisms for novel nucleases is challenging.
• Existing methods for nuclease detection can be time-consuming and costly.

Purpose of the Study:

• To develop a rapid, cost-effective 3-step screening strategy for identifying extracellular nucleases in marine organisms.
• To validate the presence and characterize a novel nuclease from a marine isolate.

Main Methods:

• A 3-step screening approach involving agar overlay assay, well-diffusion assay, and ammonium sulfate fractionation.
• Screening of approximately 500 bacterial colonies from marine environments.
• Characterization of the identified nuclease's enzymatic properties, including substrate specificity, cofactor dependency, salt tolerance, and optimal temperature.

Main Results:

• The 3-step strategy successfully identified potential nuclease-producing isolates from marine bacteria.
• Isolate SH1 exhibited significant extracellular nuclease activity.
• The characterized nuclease is a multi-subunit enzyme with both endo- and exonuclease activity, degrading DNA and RNA, requiring Mg+2, and exhibiting broad salt tolerance (80-1500 mM) and optimal activity at 37°C.

Conclusions:

• The developed 3-step strategy is effective for screening and validating extracellular nucleases from marine organisms.
• The identified nuclease from isolate SH1 possesses unique properties, suggesting potential for biotechnological applications.
• This screening approach facilitates the discovery of novel enzymes from extremophiles for further bioengineering.