Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress

Verona, Italy - 24/27 September, 2003

ISBN 88-900622-4-X

Poster Abstract - 5.31

PHOTOSYSTEM II BASED BIOSENSORS: OVEREXPRESSION OF D1 PROTEIN IN E. COLI

G. TESTONE*, D. GIANNINO*, E. PACE**, D. MARIOTTI*, M. T. GIARDI**

*) Istituto di Biologia e Biotecnologie Agrarie (IBBA), CNR, Area della Ricerca di Roma, Via Salaria Km. 29,300, 00016 Monterotondo Scalo (Roma), IT

**) Istituto di Cristallografia (IC), CNR, Area della Ricerca di Roma, Via Salaria Km. 29,300, 00016 Monterotondo Scalo (Roma), IT

psbA gene, D1 protein, Chlamydomonas reinhardtii, biosensors

The development of biosensors responds to the demand for in-situ, rapid, simple, selective and inexpensive tools for pesticides monitoring. A biosensor is defined as a analytical device, incorporating a biological or biomimetic sensing element that provides selectivity and is closely associated to a transducer system. Several currently used herbicides mostly affect the photosystem II (PSII) complex of plants and impair the light dependent phase of photosynthesis. The PSII is a multi-enzymatic chlorophyll-protein complex located in the thylakoids of algae, cyanobacteria and higher plants and it may turn useful as a component for herbicide biosensors upon extraction from the organisms. The PSII-D1 and D2 proteins together are responsible for the electron transfer to the plastoquinone in the electron cascade of photosynthesis, though D1 is the major target of herbicides. In order to construct a new class of biosensors, wild type, naturally mutated and PCR-mutagenised D1 proteins from Chlamydomonas reinhardtii were over expressed in E. coli to: a) produce bacterial liquid culture based biosensors and b) purify D1 proteins to be fixed on supports, which are necessary for new biosensor prototypes. Here, data on the efficiency of D1 overexpression in E. coli BL21-SI (gateway cloning technology), protein yield and sub cellular location, and D1/herbicide interaction assays will be presented and discussed.