Hydrolysis of RNA by transition metal complexes

Stern, Michael; Bashkin, James K.; Sall, Erik D.

doi:10.1021/ja00169a057

Cited by 120 publications

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“…The fact that the cleavage occurs upon mixing at 25°C contrasts the action of an increasing number of metal complexes that hydrolyze RNA (not DNA) over 4-20 hr at 37°C (15)(16)(17)(18). It is also interesting to note that the cleavage affords exclusively 3'-OH and 5'-OP03 ends, as found for hydrolysis mediated by restriction enzymes.…”

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confidence: 89%

“…Indeed metal centers play a significant role in the hydrolysis of phosphate esters in biology (10)(11)(12). A number of metal ion complexes have been shown to hydrolyze phosphate esters (13,14) as well as RNA with various efficiencies (15)(16)(17)(18); however, these complexes are unable to hydrolyze the phosphate diester backbone of DNA. The only reported metal-catalyzed DNA hydrolysis thus far is effected by Cu2+, Zn2+, Cd2+, or Pb2+ complexed to a polyamine ligand tethered to a DNA-binding [Ru(phen)3]2+ derivative (19), which afforded nicked (single-stranded break) pBR322 at 37°C after 5-7 hr.…”

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confidence: 99%

“…This article must therefore be hereby marked "advertisement" in accordance with 18 (20). This complex has been shown to interact with H202 to form a 1:1 adduct (20).…”

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Double-stranded cleavage of pBR322 by a diiron complex via a "hydrolytic" mechanism.

Hanson

Que

1994

Proc. Natl. Acad. Sci. U.S.A.

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Treatment of plasmid pBR322 with Fe2-(HPTB)(OH)(NO3)4(HPTB =N,N,N',N '-tetrakis(2-benzimidazolylmethyl The complex Fe2(HPTB)(OH)(NO3)4 was synthesized according to published procedures (20). This complex has been shown to interact with H202 to form a 1:1 adduct (20).Plasmid pBR322 was purchased from Promega and transformed into Escherichia coli strains JM109 and HB101 by using CaCl2 (22). The cells were grown in Luria-Bertani broth containing tetracycline, amplified with chloramphenicol, and harvested; the plasmid was purified according to published procedures (23). Purity and quantity of DNA were determined by measuring the A26o/A280 ratio, using a Hoefer fluorometer and gel electrophoresis.Linearization of pBR322 with Fe2(HPTB)(OH)(NO34. The cleavage of pBR322 by Fe2(HPTB)(OH)(NO3)4/H202 was accomplished by mixing (in order) 1 ,ul of (0.15 mg/ml) pBR322, 1 ,ul of 0.1 M Tris HCl (pH 8.0) buffer, 1,ul (0.1 mM) of Fe2(HPTB)(OH)(NO3)4, 6 ,l of H20, and 1 ,ul of 9.0 mM H202 at 25°C. Immediately after mixing, the DNA solution was subjected to electrophoresis on a 1% agarose gel in TAE (40 mM Tris acetate/i mM EDTA) (24) and stained with ethidium bromide solution at 0.5 pg/ml to observe the cleaved DNA products. The amount of DNA was quantitated by photographing the fluorescence under UV light and analyzing the band intensities with a Pharmacia densitometer. The fluorescence intensities were corrected for the different binding affinities ofethidium bromide to form I vs. forms II and III (25). Lengthening the reaction time did not affect the amounts of nicked and linearized pBR322. The linearized pBR322 was recovered from the agarose gel by electroelution ofthe excised gel fragment, followed by phenol/chloroform extraction and precipitation with 3.0 M NaOAc and ethanol (24).Control experiments with Fe(EDTA)/H202 were done under conditions identical to those of Fe2(HPTB)(OH)(NO3)4/ Abbreviations:

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Double-stranded cleavage of pBR322 by a diiron complex via a "hydrolytic" mechanism.

Hanson

Que

1994

Proc. Natl. Acad. Sci. U.S.A.

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“…In this system, the rate determining step is always the first step, in which the hydroxyl group adds to the phosphate to form a phosphorane, since the nitrophenoxide ion is such a good leaving group that forward decomposition of the phosphorane is always faster than the reverse of the first step, reopening of the phosphorane to form starting material. The cyclization/cleavage of compound 4 is convenient to follow by UV spectroscopy, although it is less closely related to the chemistry of RNA itself than are the reactions of compound 1. Several examples are known of the cleavage of RNA by Cu2+ (6), by Pb2+ (7,8), and by rare earth ions such as Eu3+ (9)(10)(11)(12). Furthermore, in ribozymes it seems clear (13,14) …”

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Effects of metal ions, including Mg2+ and lanthanides, on the cleavage of ribonucleotides and RNA model compounds.

Breslow

Huang

1991

Proc. Natl. Acad. Sci. U.S.A.

156

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The cyclization/cleavage of 3',5"-uridyluridine to form 2',3'-cycfic uridylic acid is very effectively catalyzed by Eu3', and the cyclization/cleavage of the l-pnitrophenyl phosphate ester of propane-1,2-diol also shows strong metal ion catalysis by Eu3+, Tb3+, and Yb3+. It also shows moderate catalysis by Mg2', but not by Ca2+; Zn2+ and Pb2+ are also good catalysts. Various ligands activate these reactions further, and imidazole apparently acts as an additional base catalyst. Some cyclodextrin derivatives act to bind both the substrate and the metal ion but, contrary to what is reported elsewhere, there is no strong selectivity among nucleotides that can be ascribed to cyclodextrin binding.Mechanistic studies on the cleavage of RNA (1) and of the representative dinucleotides UpU (2, 3) and ApA (4) have been described. Most ofthis work has focused on catalysis by imidazole buffers; the components imidazole and imidazolium ion are the major catalytic groups in the enzyme ribonuclease A. The preferred mechanism for cleavage of poly(U), of 3',5"-UpU (1), and of 3',5"-ApA involved protonation of the phosphate anion, followed by imidazolepromoted cyclization to form a phosphorane and then imidazole-catalyzed conversion ofthat phosphorane to the 2',3'-cyclic phosphate product characteristically formed during chemical or enzymatic cleavage of RNA (Scheme I). The evidence that ribonuclease A uses a closely related mechanism, but not the standard one found in textbooks, was pointed out, except that what is sequential in simple chemical catalysis becomes simultaneous in the enzyme (2, 3). Recently, calculated reaction paths (5) for the enzyme have supported our proposal.Although ribonuclease A does not use metal ions for catalysis, many reactions of RNA and DNA do involve metals. Thus we examined the effectiveness, and mechanism of action, of Zn2+ in the cleavage of two models for RNA (2). In one system (Scheme HA), the cyclization/cleavage of 3',5"-UpU 1 was studied; this is the same reaction in which the catalysis by imidazole buffers that led to our mechanistic proposals for the enzyme was observed (3). UpU is reallyjust a small piece of RNA. Under some conditions the slow step is the cyclization of compound 1 to form a phosphorane intermediate (2); under other conditions compound 2 is rapidly and reversibly formed, and its decomposition to form the product compound 3 is the slow step. We have shown that compound 2 is also the intermediate in the rearrangement of compound 1 to its 2',5" isomer.The second system (Scheme IIB) involves the cyclization/ cleavage of a simpler analog (4). In this system, the rate determining step is always the first step, in which the hydroxyl group adds to the phosphate to form a phosphorane, since the nitrophenoxide ion is such a good leaving group that forward decomposition of the phosphorane is always faster than the reverse of the first step, reopening of the phosphorane to form starting material. The cyclization/cleavage of compound 4 is convenient to follow by UV spectroscopy,...

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“…In this case, TCAA may enhance the hydrolytic activity by forming a Cu-TCAA complex. This mechanism may resemble the enhancement of RNA hydrolysis by several Cu(II) complexes Sigman, 1986;Stern et al, 1990;Modak et al, 1991;Burstyn and Deal, 1993;Linkletter and Chin, 1995).…”

Section: Effect Of Tcaa For the First Transition Metal Ion Catalystsmentioning

confidence: 92%

Inhibitory activity of thermal copolymers of amino acids for the metal-catalyzed hydrolysis of an RNA dinucleotide

Kawamura

Nagahama

Yao

2006

Advances in Space Research

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Hydrolysis of RNA by transition metal complexes

Cited by 120 publications

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Double-stranded cleavage of pBR322 by a diiron complex via a "hydrolytic" mechanism.

Double-stranded cleavage of pBR322 by a diiron complex via a "hydrolytic" mechanism.

Effects of metal ions, including Mg2+ and lanthanides, on the cleavage of ribonucleotides and RNA model compounds.

Inhibitory activity of thermal copolymers of amino acids for the metal-catalyzed hydrolysis of an RNA dinucleotide

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