Poster: Phytoremediation

Abs # 979: Heavy Metal Binding Properties of Chlamydomonas Cells Expressing Genes Encoding Periplasmic Metallothionein Fusion Proteins

Presenter:	Sayre, Richard T, sayre.2@osu.edu
Authors	Sayre, Richard T (A)   Traina, Samuel  (B)   Siripornadulsil, Surasak  (C)
Affiliations:	(A): Department of Plant Biology and Biophysics, Ohio State University (B): University of California, Merced (C): Biophysics Program, Ohio State University

Metallothionein (MT) is a low molecular weight, cysteine-rich metal-binding protein. MTs usually contain 61 to 62 amino acids, 20 of which are cysteine residues whose thiolate sulfur atoms serve as metal ligands. MTs can bind many kinds of heavy metals including copper, zinc, cadmium, mercury, nickel, cobalt, silver, gold, and lead. The MT protein has two distinct domains,alpha and beta. Monovalent and divalent metals interact differentially with the two structural domains. Monovalent ions, such as Cu+1 and Ag+1, preferentially bind to the beta-domain. In contrast, divalent ions, such as Cd+2 and Zn+2, bind to the alpha domain. We have constructed a fusion gene between a chicken MT-II gene and a Chlamydomonas gene encoding a plasma membrane protein to both anchor the MT-II domain on the exterior surface of the cell and to stabilize the protein. To increase the specificity of transgenic Chlamydomonas for monovalent or divalent ions, we have constructed fusion genes between the alpha or beta domain of chicken MT-II and the plasma membrane protein. Finally, to increase the heavy metal binding capacity of the transformant, we have constructed a series of polymers of chicken MT-II, the alpha domain, and/or the beta-domain, from 1-5 units in length fused to the Chlamydomonas plasma membrane protein. We show that transgenic algae expressing the synthetic genes have substantially enhanced tolerance to toxic cadmium (Cd) concentrations (100 uM). In addition, the Cd binding capacity increased 2 to 5-fold relative to wild-type cells when grown in the presence of 50 uM Cd. Sensitivity to EDTA removal of the metal ions and EXAFS studies are consistent with coordination of the metal ions by the MT-II cysteinyl sulfides.