Zn2+-dependent deoxyribozymes that form natural and unnatural RNA linkages

Kelly A. Hoadley, Whitney E. Purtha, Amanda C. Wolf, Amber Flynn-Charlebois, Scott K Silverman

Research output: Contribution to journalArticle

Abstract

We report Zn2+-dependent deoxyribozymes that ligate RNA. The DNA enzymes were identified by in vitro selection and ligate RNA with k obs up to 0.5 min-1 at 1 mM Zn2+ and 23 °C, pH 7.9, which is substantially faster than our previously reported Mg 2+-dependent deoxyribozymes. Each new Zn2+-dependent deoxyribozyme mediates the reaction of a specific nucleophile on one RNA substrate with a 2′,3′-cyclic phosphate on a second RNA substrate. Some of the Zn2+-dependent deoxyribozymes create native 3′-5′ RNA linkages (with kobs up to 0.02 min -1), whereas all of our previous Mg2+-dependent deoxyribozymes that use a 2′,3′-cyclic phosphate create non-native 2′-5′ RNA linkages. On this basis, Zn2+-dependent deoxyribozymes have promise for synthesis of native 3′-5′-linked RNA using 2′,3′-cyclic phosphate RNA substrates, although these particular Zn2+-dependent deoxyribozymes are likely not useful for this practical application. Some of the new Zn2+-dependent deoxyribozymes instead create non-native 2′-5′ linkages, just like their Mg2+ counterparts. Unexpectedly, other Zn2+- dependent deoxyribozymes synthesize one of three unnatural linkages that are formed upon the reaction of an RNA nucleophile other than a 5′-hydroxyl group. Two of these unnatural linkages are the 3′-2′ and 2′-2′ linear junctions created when the 2′-hydroxyl of the 5′-terminal guanosine of one RNA substrate attacks the 2′,3′-cyclic phosphate of the second RNA substrate. The third unnatural linkage is a branched RNA that results from attack of a specific internal 2′-hydroxyl of one RNA substrate at the 2′,3′-cyclic phosphate. When compared with the consistent creation of 2′-5′ linkages by Mg2+-dependent ligation, formation of this variety of RNA ligation products by Zn2+-dependent deoxyribozymes highlights the versatility of transition metals such as Zn2+ for mediating nucleic acid catalysis.

Original languageEnglish (US)
Pages (from-to)9217-9231
Number of pages15
JournalBiochemistry
Volume44
Issue number25
DOIs
StatePublished - Jun 28 2005

Fingerprint

Catalytic DNA
RNA
Phosphates
Substrates
Hydroxyl Radical
Nucleophiles
Ligation
Guanosine
Catalysis

ASJC Scopus subject areas

  • Biochemistry

Cite this

Hoadley, K. A., Purtha, W. E., Wolf, A. C., Flynn-Charlebois, A., & Silverman, S. K. (2005). Zn2+-dependent deoxyribozymes that form natural and unnatural RNA linkages. Biochemistry, 44(25), 9217-9231. https://doi.org/10.1021/bi050146g

Zn2+-dependent deoxyribozymes that form natural and unnatural RNA linkages. / Hoadley, Kelly A.; Purtha, Whitney E.; Wolf, Amanda C.; Flynn-Charlebois, Amber; Silverman, Scott K.

In: Biochemistry, Vol. 44, No. 25, 28.06.2005, p. 9217-9231.

Research output: Contribution to journalArticle

Hoadley, KA, Purtha, WE, Wolf, AC, Flynn-Charlebois, A & Silverman, SK 2005, 'Zn2+-dependent deoxyribozymes that form natural and unnatural RNA linkages', Biochemistry, vol. 44, no. 25, pp. 9217-9231. https://doi.org/10.1021/bi050146g
Hoadley, Kelly A. ; Purtha, Whitney E. ; Wolf, Amanda C. ; Flynn-Charlebois, Amber ; Silverman, Scott K. / Zn2+-dependent deoxyribozymes that form natural and unnatural RNA linkages. In: Biochemistry. 2005 ; Vol. 44, No. 25. pp. 9217-9231.
@article{ff4853a1539a4e9682f70d35596dc00e,
title = "Zn2+-dependent deoxyribozymes that form natural and unnatural RNA linkages",
abstract = "We report Zn2+-dependent deoxyribozymes that ligate RNA. The DNA enzymes were identified by in vitro selection and ligate RNA with k obs up to 0.5 min-1 at 1 mM Zn2+ and 23 °C, pH 7.9, which is substantially faster than our previously reported Mg 2+-dependent deoxyribozymes. Each new Zn2+-dependent deoxyribozyme mediates the reaction of a specific nucleophile on one RNA substrate with a 2′,3′-cyclic phosphate on a second RNA substrate. Some of the Zn2+-dependent deoxyribozymes create native 3′-5′ RNA linkages (with kobs up to 0.02 min -1), whereas all of our previous Mg2+-dependent deoxyribozymes that use a 2′,3′-cyclic phosphate create non-native 2′-5′ RNA linkages. On this basis, Zn2+-dependent deoxyribozymes have promise for synthesis of native 3′-5′-linked RNA using 2′,3′-cyclic phosphate RNA substrates, although these particular Zn2+-dependent deoxyribozymes are likely not useful for this practical application. Some of the new Zn2+-dependent deoxyribozymes instead create non-native 2′-5′ linkages, just like their Mg2+ counterparts. Unexpectedly, other Zn2+- dependent deoxyribozymes synthesize one of three unnatural linkages that are formed upon the reaction of an RNA nucleophile other than a 5′-hydroxyl group. Two of these unnatural linkages are the 3′-2′ and 2′-2′ linear junctions created when the 2′-hydroxyl of the 5′-terminal guanosine of one RNA substrate attacks the 2′,3′-cyclic phosphate of the second RNA substrate. The third unnatural linkage is a branched RNA that results from attack of a specific internal 2′-hydroxyl of one RNA substrate at the 2′,3′-cyclic phosphate. When compared with the consistent creation of 2′-5′ linkages by Mg2+-dependent ligation, formation of this variety of RNA ligation products by Zn2+-dependent deoxyribozymes highlights the versatility of transition metals such as Zn2+ for mediating nucleic acid catalysis.",
author = "Hoadley, {Kelly A.} and Purtha, {Whitney E.} and Wolf, {Amanda C.} and Amber Flynn-Charlebois and Silverman, {Scott K}",
year = "2005",
month = "6",
day = "28",
doi = "10.1021/bi050146g",
language = "English (US)",
volume = "44",
pages = "9217--9231",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "25",

}

TY - JOUR

T1 - Zn2+-dependent deoxyribozymes that form natural and unnatural RNA linkages

AU - Hoadley, Kelly A.

AU - Purtha, Whitney E.

AU - Wolf, Amanda C.

AU - Flynn-Charlebois, Amber

AU - Silverman, Scott K

PY - 2005/6/28

Y1 - 2005/6/28

N2 - We report Zn2+-dependent deoxyribozymes that ligate RNA. The DNA enzymes were identified by in vitro selection and ligate RNA with k obs up to 0.5 min-1 at 1 mM Zn2+ and 23 °C, pH 7.9, which is substantially faster than our previously reported Mg 2+-dependent deoxyribozymes. Each new Zn2+-dependent deoxyribozyme mediates the reaction of a specific nucleophile on one RNA substrate with a 2′,3′-cyclic phosphate on a second RNA substrate. Some of the Zn2+-dependent deoxyribozymes create native 3′-5′ RNA linkages (with kobs up to 0.02 min -1), whereas all of our previous Mg2+-dependent deoxyribozymes that use a 2′,3′-cyclic phosphate create non-native 2′-5′ RNA linkages. On this basis, Zn2+-dependent deoxyribozymes have promise for synthesis of native 3′-5′-linked RNA using 2′,3′-cyclic phosphate RNA substrates, although these particular Zn2+-dependent deoxyribozymes are likely not useful for this practical application. Some of the new Zn2+-dependent deoxyribozymes instead create non-native 2′-5′ linkages, just like their Mg2+ counterparts. Unexpectedly, other Zn2+- dependent deoxyribozymes synthesize one of three unnatural linkages that are formed upon the reaction of an RNA nucleophile other than a 5′-hydroxyl group. Two of these unnatural linkages are the 3′-2′ and 2′-2′ linear junctions created when the 2′-hydroxyl of the 5′-terminal guanosine of one RNA substrate attacks the 2′,3′-cyclic phosphate of the second RNA substrate. The third unnatural linkage is a branched RNA that results from attack of a specific internal 2′-hydroxyl of one RNA substrate at the 2′,3′-cyclic phosphate. When compared with the consistent creation of 2′-5′ linkages by Mg2+-dependent ligation, formation of this variety of RNA ligation products by Zn2+-dependent deoxyribozymes highlights the versatility of transition metals such as Zn2+ for mediating nucleic acid catalysis.

AB - We report Zn2+-dependent deoxyribozymes that ligate RNA. The DNA enzymes were identified by in vitro selection and ligate RNA with k obs up to 0.5 min-1 at 1 mM Zn2+ and 23 °C, pH 7.9, which is substantially faster than our previously reported Mg 2+-dependent deoxyribozymes. Each new Zn2+-dependent deoxyribozyme mediates the reaction of a specific nucleophile on one RNA substrate with a 2′,3′-cyclic phosphate on a second RNA substrate. Some of the Zn2+-dependent deoxyribozymes create native 3′-5′ RNA linkages (with kobs up to 0.02 min -1), whereas all of our previous Mg2+-dependent deoxyribozymes that use a 2′,3′-cyclic phosphate create non-native 2′-5′ RNA linkages. On this basis, Zn2+-dependent deoxyribozymes have promise for synthesis of native 3′-5′-linked RNA using 2′,3′-cyclic phosphate RNA substrates, although these particular Zn2+-dependent deoxyribozymes are likely not useful for this practical application. Some of the new Zn2+-dependent deoxyribozymes instead create non-native 2′-5′ linkages, just like their Mg2+ counterparts. Unexpectedly, other Zn2+- dependent deoxyribozymes synthesize one of three unnatural linkages that are formed upon the reaction of an RNA nucleophile other than a 5′-hydroxyl group. Two of these unnatural linkages are the 3′-2′ and 2′-2′ linear junctions created when the 2′-hydroxyl of the 5′-terminal guanosine of one RNA substrate attacks the 2′,3′-cyclic phosphate of the second RNA substrate. The third unnatural linkage is a branched RNA that results from attack of a specific internal 2′-hydroxyl of one RNA substrate at the 2′,3′-cyclic phosphate. When compared with the consistent creation of 2′-5′ linkages by Mg2+-dependent ligation, formation of this variety of RNA ligation products by Zn2+-dependent deoxyribozymes highlights the versatility of transition metals such as Zn2+ for mediating nucleic acid catalysis.

UR - http://www.scopus.com/inward/record.url?scp=21744451987&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=21744451987&partnerID=8YFLogxK

U2 - 10.1021/bi050146g

DO - 10.1021/bi050146g

M3 - Article

C2 - 15966746

AN - SCOPUS:21744451987

VL - 44

SP - 9217

EP - 9231

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 25

ER -