Babic I, Andrew S E, Jirik F R
Biomedical Research Center, University of British Columbia, Vancouver, Canada.
Mutat Res. 1996 Nov 11;372(1):87-96. doi: 10.1016/S0027-5107(96)00170-4.
An optical biosensor was used to monitor interactions between the Escherichia coli DNA mismatch repair molecule MutS and various immobilized oligonucleotides. While associating poorly with single-stranded DNA, MutS was capable of rapid association/dissociation from homoduplex DNA. The interaction of MutS with oligonucleotide 30-mers containing single site mismatches demonstrated that during the dissociation phase, MutS binding was greatest to a G-G mismatch, followed by G-T > A-A > C-T, A-C. Binding to A-G, T-T and C-C mispairs was marginally higher than that seen between MutS and homoduplex DNA. The ability of MutS to interact with 30-mers containing alkylated bases was also tested. While binding to O6-methyl-G-C, or to O4-methyl-T-A base pairs was similar to that of homoduplex DNA, strong binding was seen to a O6-methyl-G-T mispair. O4-methyl-T-G, however, was poorly recognized by MutS, with relative binding affinity similar to homoduplex DNA, predicting poor in vivo recognition of O4-methyl-T-G by MutS. Interestingly, MutS demonstrated a relatively high affinity for an 1,N6-etheno-A-T containing homoduplex. Thus, in allowing rapid evaluation of interactions between such molecules, the biosensor will be useful to structure-function analyses.
一种光学生物传感器被用于监测大肠杆菌DNA错配修复分子MutS与各种固定化寡核苷酸之间的相互作用。虽然MutS与单链DNA的结合较差，但它能够与同型双链DNA快速结合/解离。MutS与含有单个位点错配的30聚体寡核苷酸的相互作用表明，在解离阶段，MutS对G-G错配的结合最强，其次是G-T > A-A > C-T、A-C。与A-G、T-T和C-C错配的结合略高于MutS与同型双链DNA之间的结合。还测试了MutS与含有烷基化碱基的30聚体相互作用的能力。虽然与O6-甲基-G-C或O4-甲基-T-A碱基对的结合与同型双链DNA相似，但对O6-甲基-G-T错配有强烈结合。然而，MutS对O4-甲基-T-G的识别较差，其相对结合亲和力与同型双链DNA相似，这预示着MutS在体内对O4-甲基-T-G的识别较差。有趣的是，MutS对含有1,N6-乙烯基-A-T的同型双链显示出相对较高的亲和力。因此，这种生物传感器能够快速评估此类分子之间的相互作用，将有助于结构-功能分析。
