1SKM

HhaI methyltransferase in complex with DNA containing an abasic south carbocyclic sugar at its target site

Summary for 1SKM

• Entry DOI: 10.2210/pdb1skm/pdb
• Related: 1FJX 1HMY 1MHT 9MHT
• Descriptor: 5'-D(*TP*CP*CP*AP*TP*GP*CP*GP*CP*TP*GP*AP*C)-3', 5'-D(*T*GP*TP*CP*AP*GP*(HCX)P*GP*CP*AP*TP*GP*G)-3', Modification methylase HhaI, ... (5 entities in total)
• Functional Keywords: protein-dna complex containing constrained abasic unnatural base, transferase-dna complex, transferase/dna
• Biological source: Haemophilus haemolyticus
• Total number of polymer chains: 3
• Total formula weight: 45246.74

Authors

Horton, J.R.,Ratner, G.,Banavali, N.,Huang, N.,Marquez, V.E.,MacKerell, A.D.,Cheng, X. (deposition date: 2004-03-05, release date: 2004-08-24, Last modification date: 2024-02-14)

Primary citation

Horton, J.R.,Ratner, G.,Banavali, N.K.,Huang, N.,Choi, Y.,Maier, M.A.,Marquez, V.E.,MacKerell, A.D.,Cheng, X.
Caught in the act: visualization of an intermediate in the DNA base-flipping pathway induced by HhaI methyltransferase
Nucleic Acids Res., 32:3877-3886, 2004

PubMed 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.

PubMed: 15273274
DOI: 10.1093/nar/gkh701

Experimental method

X-RAY DIFFRACTION (2.2 Å)