Altering the Double-Stranded DNA Specificity of the bZIP Domain of Zta with Site-Directed Mutagenesis at N182

被引：1
作者：
Ray, Sreejana [1 ]
Tillo, Desiree [1 ,2 ]
Assad, Nima [1 ]
Ufot, Aniekanabasi [1 ]
Porollo, Aleksey [3 ,4 ]
Durell, Stewart R. [5 ]
Vinson, Charles [1 ]
机构：
[1] NCI, Lab Metab, NIH, Bethesda, MD 20892 USA
[2] NCI, Canc Genet Branch, NIH, Bethesda, MD 20892 USA
[3] Cincinnati Childrens Hosp Med Ctr, Ctr Autoimmune Genom & Etiol, Div Biomed Informat, Cincinnati, OH 45229 USA
[4] Univ Cincinnati, Dept Pediat, Coll Med, Cincinnati, OH 45267 USA
[5] NCI, Lab Cell Biol, NIH, Bethesda, MD 20892 USA
来源：
ACS OMEGA | 2022年 / 7卷 / 01期
关键词：
TRANSCRIPTION-FACTOR; BASIC REGION; CRYSTAL-STRUCTURE; STRUCTURAL BASIS; DIMERIZATION SPECIFICITY; NEGATIVE DESIGN; PROTEIN-BINDING; RECOGNITION; AP-1; MICROARRAYS;
D O I：
10.1021/acsomega.1c04148
中图分类号：
O6 [化学];
学科分类号：
0703 ;
摘要：
Zta, the Epstein-Barr virus bZIP transcription factor (TF), binds both unmethylated and methylated double-stranded DNA (dsDNA) in a sequence-specific manner. We studied the contribution of a conserved asparagine (N182) to sequence-specific dsDNA binding to four types of dsDNA: (i) dsDNA with cytosine in both strands ((DNA(C vertical bar C)), (ii, iii) dsDNA with 5-methylcytosine (5mC, M) or 5-hydroxymethylcytosine (5hmC, H) in one strand and cytosine in the second strand ((DNA(5mC vertical bar C) and DNA(5hmC vertical bar C)), and (iv) dsDNA with methylated cytosine in both strands in all CG dinucleotides ((DNA(5mCG)). We replaced asparagine with five similarly sized amino acids (glutamine (Q), serine (S), threonine (T), isoleucine (I), or valine (V)) and used protein binding microarrays to evaluate sequence-specific dsDNA binding. Zta preferentially binds the pseudo-palindrome TRE (AP1) motif (T(-4)G(-3)A(-2G)/ (0)(C)T(2)C(3)A(4)). Zta (N182Q) changes binding to A3 in only one half-site. Zta(N182S) changes binding to G3 in one or both halves of the motif. Zta(N182S) and Zta(N182Q) have 34- and 17-fold weaker median dsDNA binding, respectively. Zta(N182V) and Zta(N182I) have increased binding to dsDNA(5mC|C). Molecular dynamics simulations rationalize some of these results, identifying hydrogen bonds between glutamine and A(3), but do not reveal why serine preferentially binds G(3), suggesting that entropic interactions may mediate this new binding specificity.
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收藏
页码：129 / 139
页数：11
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