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Norwegian - Danish collaboration uncovers proton channel in a bacterial enzyme

A scientific collaboration between researchers from the University of Oslo and Aarhus University has revealed an unexpected channel for removal of protons in the enzyme isatin hydrolase.

The Danish research team behind the new results (from left): Jan K. Jensen, Michael Etzerodt, Anette Kjems and Bjarne Jochimsen (photo: Lisbeth Heilesen)
The Norwegian research team: Kaare Bjerregaard-Andersen, J. Preben Morth and Theis Sommer (photo: Oslo University)
Identification of two entrances to the active site, a combined water and proton channel and a substrate and product channel (figure: Kaare Bjerregaard-Andersen)

Isatin hydrolase, which is involved in the breakdown of the plant hormone indole-3-acetic acid, has been identified in a wide variety of bacterial isolates from soil, water, plants and the human gut. The first identification was done with the soil bacterium Bradyrhizobium japonicum, which forms nitrogen fixing root nodules on the host plant soybean. As the marine bacterium Labrenzia aggregata possesses two genes encoding isatin hydrolase, the researchers chose to study the enzymes from this organism in detail.

PhD student Kaare Bjerregaard-Andersen succeeded in obtaining crystals of the two isatin hydrolases as well as determining the three-dimensional structure. The dimeric protein reveals a new protein-fold for a metal-dependent (here Mn++) hydrolytic enzyme, in which the two subunits exhibit overlapping domains. More interesting is the identification of two entrances to the active site, a combined water and proton channel and a substrate and product channel, as shown in the figure. The basis for the identification of a water/proton channel has been crystals, where an analogue of the product (thioisatin)  is trapped in the active site. Isatin hydrolase has a pH optimum of 9, so removal of the protons formed in the reaction is needed.

In addition to isatin being an intermediate in the degradation of indole-3-acetic acid, isatin is present in low concentrations in the blood of humans, and it is likely to be a degradation product of neurotransmitters. The level of isatin is shown to be elevated in patients with stress, and patients suffering from Parkinson's disease.  The Norwegian-Danish research team has conducted preliminary experiments aimed at using isatin hydrolase in a simple enzymatic screening test for the determination of isatin in blood and urine.


Kaare Bjerregaard-Andersen is the first Danish student to obtain a double PhD degree, as part of an agreement between the University of Oslo and the University of Aarhus. Today, 30 June 2014, Kaare defends his PhD dissertation at the University of Oslo.

Link to the article in the Journal of Biological Chemistry: A proton wire and water channel revealed in the crystal structure of isatin hydrolase


More information

Associate Professor Bjarne Jochimsen
Department of Molecular Biology and Genetics
Aarhus University, Denmark
bjoc@mb.au.dk - +45 20275983