Caught in the act

Visualization of an intermediate in the DNA base-flipping pathway induced by Hhal methyltransferase

John R. Horton, Gary Ratner, Nilesh K. Banavali, Niu Huang, Yongseok Choi, Martin A. Maier, Victor E. Marquez, Alexander D. MacKerell, Xiaodong Cheng

Research output: Contribution to journalArticle

41 Citations (Scopus)

Abstract

Rotation of a DNA or RNA nucleotide out of the double helix and into a protein pocket ('base flipping') is a mechanistic feature common to some DNA/RNA-binding proteins. Here, we report the structure of HhaI methyltransferase in complex with DNA containing a south-constrained abasic carbocyclic sugar at the target site in the presence of the methyl donor byproduct AdoHcy. Unexpectedly, the locked south pseudosugar appears to be trapped in the middle of the flipping pathway via the DNA major groove, held in place primarily through Van der Waals contacts with a set of invariant amino acids. Molecular dynamics simulations indicate that the structural stabilization observed with the south-constrained pseudosugar will not occur with a north-constrained pseudosugar, which explains its lowered binding affinity. Moreover, comparison of structural transitions of the sugar and phosphodiester backbone observed during computational studies of base flipping in the M.HhaI-DNA-AdoHcy ternary complex indicate that the south-constrained pseudosugar induces a conformation on the phosphodiester backbone that corresponds to that of a discrete intermediate of the base-flipping pathway. As previous crystal structures of M.HhaI ternary complex with DNA displayed the flipped sugar moiety in the antipodal north conformation, we suggest that conversion of the sugar pucker from south to north beyond the middle of the pathway is an essential part of the mechanism through which flipping must proceed to reach its final destination. We also discuss the possibility of the south-constrained pseudosugar mimicking a transition state in the phosphodiester and sugar moieties that occurs during DNA base flipping in the presence of M.HhaI.

Original languageEnglish
Pages (from-to)3877-3886
Number of pages10
JournalNucleic Acids Research
Volume32
Issue number13
DOIs
Publication statusPublished - 2004 Sep 21
Externally publishedYes

Fingerprint

Carbasugars
Methyltransferases
Visualization
Sugars
DNA
Conformations
RNA-Binding Proteins
DNA-Binding Proteins
Molecular Dynamics Simulation
Nucleotides
Byproducts
Molecular dynamics
RNA
Stabilization
Crystal structure
Amino Acids
Computer simulation

ASJC Scopus subject areas

  • Genetics

Cite this

Caught in the act : Visualization of an intermediate in the DNA base-flipping pathway induced by Hhal methyltransferase. / Horton, John R.; Ratner, Gary; Banavali, Nilesh K.; Huang, Niu; Choi, Yongseok; Maier, Martin A.; Marquez, Victor E.; MacKerell, Alexander D.; Cheng, Xiaodong.

In: Nucleic Acids Research, Vol. 32, No. 13, 21.09.2004, p. 3877-3886.

Research output: Contribution to journalArticle

Horton, JR, Ratner, G, Banavali, NK, Huang, N, Choi, Y, Maier, MA, Marquez, VE, MacKerell, AD & Cheng, X 2004, 'Caught in the act: Visualization of an intermediate in the DNA base-flipping pathway induced by Hhal methyltransferase', Nucleic Acids Research, vol. 32, no. 13, pp. 3877-3886. https://doi.org/10.1093/nar/gkh701
Horton, John R. ; Ratner, Gary ; Banavali, Nilesh K. ; Huang, Niu ; Choi, Yongseok ; Maier, Martin A. ; Marquez, Victor E. ; MacKerell, Alexander D. ; Cheng, Xiaodong. / Caught in the act : Visualization of an intermediate in the DNA base-flipping pathway induced by Hhal methyltransferase. In: Nucleic Acids Research. 2004 ; Vol. 32, No. 13. pp. 3877-3886.
@article{fca9cf57fa5a42179b0de8e931dd059e,
title = "Caught in the act: Visualization of an intermediate in the DNA base-flipping pathway induced by Hhal methyltransferase",
abstract = "Rotation of a DNA or RNA nucleotide out of the double helix and into a protein pocket ('base flipping') is a mechanistic feature common to some DNA/RNA-binding proteins. Here, we report the structure of HhaI methyltransferase in complex with DNA containing a south-constrained abasic carbocyclic sugar at the target site in the presence of the methyl donor byproduct AdoHcy. Unexpectedly, the locked south pseudosugar appears to be trapped in the middle of the flipping pathway via the DNA major groove, held in place primarily through Van der Waals contacts with a set of invariant amino acids. Molecular dynamics simulations indicate that the structural stabilization observed with the south-constrained pseudosugar will not occur with a north-constrained pseudosugar, which explains its lowered binding affinity. Moreover, comparison of structural transitions of the sugar and phosphodiester backbone observed during computational studies of base flipping in the M.HhaI-DNA-AdoHcy ternary complex indicate that the south-constrained pseudosugar induces a conformation on the phosphodiester backbone that corresponds to that of a discrete intermediate of the base-flipping pathway. As previous crystal structures of M.HhaI ternary complex with DNA displayed the flipped sugar moiety in the antipodal north conformation, we suggest that conversion of the sugar pucker from south to north beyond the middle of the pathway is an essential part of the mechanism through which flipping must proceed to reach its final destination. We also discuss the possibility of the south-constrained pseudosugar mimicking a transition state in the phosphodiester and sugar moieties that occurs during DNA base flipping in the presence of M.HhaI.",
author = "Horton, {John R.} and Gary Ratner and Banavali, {Nilesh K.} and Niu Huang and Yongseok Choi and Maier, {Martin A.} and Marquez, {Victor E.} and MacKerell, {Alexander D.} and Xiaodong Cheng",
year = "2004",
month = "9",
day = "21",
doi = "10.1093/nar/gkh701",
language = "English",
volume = "32",
pages = "3877--3886",
journal = "The BMJ",
issn = "0730-6512",
publisher = "Kluwer Academic Publishers",
number = "13",

}

TY - JOUR

T1 - Caught in the act

T2 - Visualization of an intermediate in the DNA base-flipping pathway induced by Hhal methyltransferase

AU - Horton, John R.

AU - Ratner, Gary

AU - Banavali, Nilesh K.

AU - Huang, Niu

AU - Choi, Yongseok

AU - Maier, Martin A.

AU - Marquez, Victor E.

AU - MacKerell, Alexander D.

AU - Cheng, Xiaodong

PY - 2004/9/21

Y1 - 2004/9/21

N2 - Rotation of a DNA or RNA nucleotide out of the double helix and into a protein pocket ('base flipping') is a mechanistic feature common to some DNA/RNA-binding proteins. Here, we report the structure of HhaI methyltransferase in complex with DNA containing a south-constrained abasic carbocyclic sugar at the target site in the presence of the methyl donor byproduct AdoHcy. Unexpectedly, the locked south pseudosugar appears to be trapped in the middle of the flipping pathway via the DNA major groove, held in place primarily through Van der Waals contacts with a set of invariant amino acids. Molecular dynamics simulations indicate that the structural stabilization observed with the south-constrained pseudosugar will not occur with a north-constrained pseudosugar, which explains its lowered binding affinity. Moreover, comparison of structural transitions of the sugar and phosphodiester backbone observed during computational studies of base flipping in the M.HhaI-DNA-AdoHcy ternary complex indicate that the south-constrained pseudosugar induces a conformation on the phosphodiester backbone that corresponds to that of a discrete intermediate of the base-flipping pathway. As previous crystal structures of M.HhaI ternary complex with DNA displayed the flipped sugar moiety in the antipodal north conformation, we suggest that conversion of the sugar pucker from south to north beyond the middle of the pathway is an essential part of the mechanism through which flipping must proceed to reach its final destination. We also discuss the possibility of the south-constrained pseudosugar mimicking a transition state in the phosphodiester and sugar moieties that occurs during DNA base flipping in the presence of M.HhaI.

AB - Rotation of a DNA or RNA nucleotide out of the double helix and into a protein pocket ('base flipping') is a mechanistic feature common to some DNA/RNA-binding proteins. Here, we report the structure of HhaI methyltransferase in complex with DNA containing a south-constrained abasic carbocyclic sugar at the target site in the presence of the methyl donor byproduct AdoHcy. Unexpectedly, the locked south pseudosugar appears to be trapped in the middle of the flipping pathway via the DNA major groove, held in place primarily through Van der Waals contacts with a set of invariant amino acids. Molecular dynamics simulations indicate that the structural stabilization observed with the south-constrained pseudosugar will not occur with a north-constrained pseudosugar, which explains its lowered binding affinity. Moreover, comparison of structural transitions of the sugar and phosphodiester backbone observed during computational studies of base flipping in the M.HhaI-DNA-AdoHcy ternary complex indicate that the south-constrained pseudosugar induces a conformation on the phosphodiester backbone that corresponds to that of a discrete intermediate of the base-flipping pathway. As previous crystal structures of M.HhaI ternary complex with DNA displayed the flipped sugar moiety in the antipodal north conformation, we suggest that conversion of the sugar pucker from south to north beyond the middle of the pathway is an essential part of the mechanism through which flipping must proceed to reach its final destination. We also discuss the possibility of the south-constrained pseudosugar mimicking a transition state in the phosphodiester and sugar moieties that occurs during DNA base flipping in the presence of M.HhaI.

UR - http://www.scopus.com/inward/record.url?scp=3242714355&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=3242714355&partnerID=8YFLogxK

U2 - 10.1093/nar/gkh701

DO - 10.1093/nar/gkh701

M3 - Article

VL - 32

SP - 3877

EP - 3886

JO - The BMJ

JF - The BMJ

SN - 0730-6512

IS - 13

ER -