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Heavy and light chain variable single domains of an anti-DNA binding antibody hydrolyze both double- and single-stranded DNAs without sequence specificity.

Authors
Kim, YR; Kim, JS; Lee, SH; Lee, WR; Sohn, JN; Chung, YC; Shim, HK; Lee, SC; Kwon, MH; Kim, YS
Citation
The Journal of biological chemistry, 281(22):15287-15295, 2006
Journal Title
The Journal of biological chemistry
ISSN
0021-92581083-351X
Abstract
Anti-DNA antibodies (Abs) are of biomedical interest because they are associated with autoimmune diseases in human and mice. Previously we isolated an anti-DNA monoclonal Ab 3D8 from an autoimmune-prone MRL-lpr/lpr mouse. Here we have characterized DNA binding kinetics and hydrolyzing activities of the recombinant single chain variable fragment (scFv) and the single variable domains of heavy chain (VH) and light chain (VL) using various single-stranded (ss) and double-stranded (ds) DNA substrates. All the Abs bound to both ds- and ssDNAs without significant preferential sequence specificity showing scFv higher affinities (KD = approximately 17-74 nm) than VH (KD = approximately 2.4-8.4 microm) and VL (KD = approximately 3.2-72 microm), and efficiently hydrolyzed both ds- and ssDNAs without sequence specificity in a Mg2+-dependent manner, except for the poor activity of 3D8 scFv for ss-(dT)40. Elucidated crystal structure-based His to Ala mutations on the complementarity determining regions of VH (His-H35 --> Ala) and/or VL (His-L94 --> Ala) of 3D8 scFv significantly inhibited the catalytic activities, indicating that the His residues are involved in the catalytic mechanism of 3D8 scFv. However, the DNA hydrolyzing activities of single domain VH and VL were not affected by the mutations, indicative of their different catalytic mechanisms from that of 3D8 scFv. Our results demonstrate single domain Abs with DNase activities for the first time, which might provide new insights into substrate recognition and catalytic mechanisms of anti-DNA Abs.
MeSH terms
Amino Acid SubstitutionAnimalsAntibodies, Antinuclear/chemistry*Antibodies, Antinuclear/geneticsAntibodies, Antinuclear/metabolism*Base SequenceCrystallography, X-RayDNA/geneticsDNA/metabolism*DNA, Single-Stranded/geneticsDNA, Single-Stranded/metabolism*HumansHydrolysisImmunoglobulin Fragments/chemistryImmunoglobulin Fragments/geneticsImmunoglobulin Fragments/metabolismImmunoglobulin Heavy Chains/chemistryImmunoglobulin Heavy Chains/geneticsImmunoglobulin Heavy Chains/metabolismImmunoglobulin Light Chains/chemistryImmunoglobulin Light Chains/geneticsImmunoglobulin Light Chains/metabolismKineticsMiceMice, Inbred MRL lprModels, MolecularMutagenesis, Site-DirectedProtein ConformationProtein Structure, TertiaryRecombinant Proteins/chemistryRecombinant Proteins/geneticsRecombinant Proteins/metabolism
DOI
10.1074/jbc.M600937200
PMID
16551636
Appears in Collections:
Journal Papers > School of Medicine / Graduate School of Medicine > Microbiology
AJOU Authors
권, 명희
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