Previous work has established that a pool of accessory proteins (called Hyp proteins) are needed for the synthesis and insertion of the NiFe(CN)2CO cofactor during maturation of the FHL complex. It is possible to purify the whole complex or individual sub-complexes, but the order of events during assembly is not understood. For example, the function of the additional accessory protein HycH remains unknown. HycH interacts with the large hydrogenase subunit HycE during assembly of the complex, but it is not a structural subunit of the FHL complex. Also, the interactions of the formate DH with the hydrogenase and the iron sulfur cluster proteins are still uncharacterized.
Understanding FHL assembly will be of great value for the biotechnological exploitation of its hydrogen-producing capability. For this reason we plan to characterise the interaction of HycH with HycE. We also want to characterize the interplay between the other subunits. Therefore, we plan to make structural investigations e.g. crystallisation and interactions studies and will determine the influence on activity and stability of the FHL complex.