An aeroplysinin-1 specific nitrile hydratase isolated from the marine sponge Aplysina cavernicola

• Published in: Marine drugs Vol. 11; no. 8; pp. 3046 - 3067
• Main Authors: Lipowicz, Bartosz, Hanekop, Nils, Schmitt, Lutz, Proksch, Peter
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 21.08.2013; MDPI
• Subjects: Acetonitriles - metabolism; alkaloids; Animals; Aplysina cavernicola; biotransformation; Cobalt - chemistry; Cyclohexenes - metabolism; Hydro-Lyases - chemistry; Hydro-Lyases - isolation & purification; Hydro-Lyases - metabolism; Hydrogen-Ion Concentration; Manganese - chemistry; Mass Spectrometry; Mediterranean Sea; Nickel - chemistry; nitrile hydratase; Porifera - chemistry; sponges; Substrate Specificity; Temperature; Mediterranean Sea
• ISSN: 1660-3397; 1660-3397
• DOI: 10.3390/md11083046

A nitrile hydratase (NHase) that specifically accepts the nitrile aeroplysinin-1 (1) as a substrate and converts it into the dienone amide verongiaquinol (7) was isolated, partially purified and characterized from the Mediterranean sponge Aplysina cavernicola; although it is currently not known whether the enzyme is of sponge origin or produced by its symbiotic microorganisms. The formation of aeroplysinin-1 and of the corresponding dienone amide is part of the chemical defence system of A. cavernicola. The latter two compounds that show strong antibiotic activity originate from brominated isoxazoline alkaloids that are thought to protect the sponges from invasion of bacterial pathogens. The sponge was shown to contain at least two NHases as two excised protein bands from a non denaturating Blue Native gel showed nitrile hydratase activity, which was not observed for control samples. The enzymes were shown to be manganese dependent, although cobalt and nickel ions were also able to recover the activity of the nitrile hydratases. The temperature and pH optimum of the studied enzymes were found at 41 °C and pH 7.8. The enzymes showed high substrate specificity towards the physiological substrate aeroplysinin-1 (1) since none of the substrate analogues that were prepared either by partial or by total synthesis were converted in an in vitro assay. Moreover de-novo sequencing by mass spectrometry was employed to obtain information about the primary structure of the studied NHases, which did not reveal any homology to known NHases.