Mercury(II) binding to s⁴U in E.coli tRNA^Val

Abstract

The accessibility of the s(4)U base in native tRNA(Val) from E.coli was monitored by studying the binding of various mercurials. The relative binding order HgBr(2)[unk]HgCl(2)>>CH(3)HgOAc[unk]CH(3)HgCl[unk]PCMB parallels approximately the steric requirements of linear HgX(2) or RHgX compounds for S(N)2 displacement by sulfur, although other factors are operative. Para-chloromercuri-benzoate (PCMB) does not bind the thiolated nucleotide unless the tertiary structure of the tRNA is opened up by removal of Mg(2+) ions and heating to 40 degrees . Under these conditions, equilibrium dialysis measurements using (14)C-labeled PCMB showed one binding site (n = 0.93) with an association constant, K(1), of 9 x 10(4)M(-1).

Tetrakis(acetoxymercuri)methane: a polymetallic reagent for labeling sulfur in nucleic acids.

“…1) has four mercury atoms bonded to a central carbon producing a compact structure (4) having 326 electrons in a sphere of -3.5 A radius. Mercury is known to bind strongly and selectively to the sulfur of 4-thiouridine residues in various Escherichia coli tRNAs (5)(6)(7)(8) and to poly[d(A-s4T)] (9), poly(s2C), and poly(s2U) (10). As demonstrated by the present work, TAMM is soluble at neutral pH in the presence of N,N-dimethyl-2-aminoethanethiol.…”

“…Titrations of E. coli tRNAval with the reagent indicate that it binds selectively to the thiolated base s4U, as has been observed for other mercury compounds (5)(6)(7)(8)(9)(10). In view of the known reactivity of mercury reagents with other nucleotides (19), the observed selectivity for 4-thiouridine of TAMM and other mercury compounds probably results from the presence of a high chloride ion concentration in the buffers used.…”

“…Sa) produced a change in the 4-thiouridine chromophore indicative of metal binding (5). The titration showed an isosbestic point and was complete at a mole ratio of TAMM to tRNA of 1, indicating strong, selective binding to the 4-thiouridine residue.…”

Platinum binds selectively to phosphorothioate groups in mono- and polynucleotides: a general method for heavy metal staining of specific nucleotides.

“…The other potential donor atoms of nucleic acids all have more class a (12), or "hard" (13), character compared to sulfur. The selectivity of heavy metals for sulfur in a polynucleotide has been previously demonstrated by the specific binding of mercury to the 4-thiouridine residues of several Escherichia coli tRNAs (14)(15)(16) and the thiolated nucleosides of polyd(A-s4T) (17), poly(s2C), and poly(s2U) (18). The present report describes the platinum binding properties of the phosphorothioate group in mono-and polynucleotides.…”

The reaction of an organomercury compound with a nucleoside phosphorothioate

“…If such a reaction takes place, the obvrous criterion in deciding whether rt wrll be quantitative under given crrcumstances is the equihbrium constant. Although K, values can be found in the literature for the reactions of RHgX compounds with several thro-analogs of nucleic acid bases [7], nucleosrdes and polynucleotrdes [8], and with s4U-containing tRNAs [9,10], no such data are available for reactrons of heavy metal compounds with nucleoside phosphorothroates.…”