Terminal tagging of full-length proteins for enhanced capturing by biological nanopores

纳米孔 终端(电信) 纳米技术 纳米孔测序 生物物理学 计算生物学 化学 生物 计算机科学 材料科学 生物化学 电信 DNA DNA测序
作者
Zhuoyu Zhang,Dylan Bloch,Luning Yu,Yu Chen,Xinqi Kang,Amr Makhamreh,Joshua C. Foster,Giovanni Maglia,Min Chen,Meni Wanunu
出处
期刊:Biophysical Journal [Elsevier BV]
卷期号:123 (3): 149a-149a
标识
DOI:10.1016/j.bpj.2023.11.1024
摘要

Having demonstrated protein fingerprinting and a strong potential for single-molecule protein sequencing, nanopores have found success in a broad range of protein translocation experiments in the past years. Most analytes tested thus far have been engineered proteins or peptide analytes, due to such limitations of native proteins as non-uniform electric charge along the chain and heterogeneity of conformation of the terminal motifs, both affecting the efficiency of capture and threading of the analyte. While this can be mitigated by engineering of the nanopore, a universal solution that normalizes sample preparation for protein analytes is still being sought by researchers in the field. Here, we show enhanced translocation of native proteins through biological nanopores by chemically tagging the N-termini of proteins with a densely charged motif such as DNA and peptide oligomers. The major challenge of such tagging chemistry is the off-target modification on the primary amines of lysine sidechains. We tested various tagging chemistry, and as expected, nanopores are susceptible to clogging from those products with lysine sidechain off-target tagging. Thus, we tuned our method to reduce such effect. We compared the capture rate and translocation signal of pristine and tagged native proteins, showing an enhancement in capture and translocation. We also compared the translocation of the tagged and the engineered versions of a protein for evidence of successful translocation. We tested several native proteins of distinct origin to show the universality of our method. Our method is generalizable to diverse architectures for protein translocation through nanopores, which will accelerate the implementation of the potential of nanopore in proteomics and unlock the reservoir of native proteins from organisms to researchers in this field.
最长约 10秒,即可获得该文献文件

科研通智能强力驱动
Strongly Powered by AbleSci AI
更新
PDF的下载单位、IP信息已删除 (2025-6-4)

科研通是完全免费的文献互助平台,具备全网最快的应助速度,最高的求助完成率。 对每一个文献求助,科研通都将尽心尽力,给求助人一个满意的交代。
实时播报
量子星尘发布了新的文献求助10
刚刚
mawenyu完成签到,获得积分10
1秒前
17完成签到,获得积分20
1秒前
高大的水壶完成签到,获得积分10
2秒前
英俊的铭应助wellyou采纳,获得10
4秒前
风中的向卉完成签到 ,获得积分10
7秒前
Mp4完成签到 ,获得积分10
7秒前
凌兰完成签到 ,获得积分10
7秒前
plain完成签到,获得积分10
8秒前
陌上花开完成签到,获得积分10
9秒前
10秒前
fg2477完成签到,获得积分10
11秒前
忙碌的数学人完成签到,获得积分10
11秒前
情怀应助Engen采纳,获得10
11秒前
HJJHJH完成签到,获得积分10
13秒前
Bob发布了新的文献求助10
14秒前
15秒前
16秒前
HJJHJH发布了新的文献求助50
17秒前
JW完成签到,获得积分10
17秒前
wanci应助张参采纳,获得10
18秒前
谦让的西装完成签到 ,获得积分10
19秒前
李演员完成签到,获得积分10
20秒前
fei菲飞完成签到,获得积分10
20秒前
22秒前
Zhaowx完成签到,获得积分10
22秒前
Theprisoners完成签到,获得积分0
22秒前
木子发布了新的文献求助30
22秒前
22秒前
下课了吧完成签到,获得积分10
23秒前
丘比特应助xialuoke采纳,获得10
24秒前
zgt01发布了新的文献求助10
26秒前
linfordlu完成签到,获得积分0
26秒前
清浅发布了新的文献求助10
27秒前
风趣的涵柏完成签到,获得积分10
28秒前
30秒前
Chen完成签到 ,获得积分10
31秒前
32秒前
木樨完成签到,获得积分10
33秒前
科研顺利完成签到,获得积分10
34秒前
高分求助中
【提示信息,请勿应助】关于scihub 10000
Les Mantodea de Guyane: Insecta, Polyneoptera [The Mantids of French Guiana] 3000
徐淮辽南地区新元古代叠层石及生物地层 3000
The Mother of All Tableaux: Order, Equivalence, and Geometry in the Large-scale Structure of Optimality Theory 3000
Handbook of Industrial Diamonds.Vol2 1100
Global Eyelash Assessment scale (GEA) 1000
Picture Books with Same-sex Parented Families: Unintentional Censorship 550
热门求助领域 (近24小时)
化学 材料科学 医学 生物 工程类 有机化学 生物化学 物理 内科学 纳米技术 计算机科学 化学工程 复合材料 遗传学 基因 物理化学 催化作用 冶金 细胞生物学 免疫学
热门帖子
关注 科研通微信公众号,转发送积分 4038235
求助须知:如何正确求助?哪些是违规求助? 3575992
关于积分的说明 11374009
捐赠科研通 3305760
什么是DOI,文献DOI怎么找? 1819276
邀请新用户注册赠送积分活动 892662
科研通“疑难数据库(出版商)”最低求助积分说明 815022