The metal binding properties of the CCCH motif of the (5)oS ribosomal protein L(3)6 from Thermus thermophilus

The TthL36 protein of the 50S ribosomal proteins from Thermus thermophilus has been found to contain the rare C(Xaa)(2)C(Xaa)(12)C(Xaa)(4)H (CCCH) seq uence motif, a zinc finger binding motif, which for other zinc finger prote ins is known to cleave RNA hairpins. In order to investigate the metal-bind ing properties of this T. thermophilus TthL36 protein, the core 26-mer poly peptide containing this CCCH motif was prepared by solid-phase peptide synt hesis methods using Fmoc chemistry, purified by preparative RP-HPLC and cha racterized by circular dichroism, high-performance capillary zone electroph oresis and electrospray ionization mass spectrometry. Reaction of the aceta midomethyl (Acm)-protected polypeptide with iodine under acidic conditions resulted in the formation of the fully deprotected polypeptide. Of interest , the results demonstrate that the standard Acm-deprotection method with th e synthetic TthL36 polypeptide using mercuric acetate in the presence of a large excess of 2-mercaptoethanol resulted in preferential formation of a v ery stable mercuro-polypeptide complex. The properties of the Acm-deprotect ed polypeptide in the presence of different metal ions were also investigat ed by spectroscopic methods. The results confirm that this TthL36 polypepti de containing the CCCH motif binds metal ions with different affinities, na mely in the order Co(II)>Hg(II)>Zn(II).