Electron Microscopic Detection of Single Membrane Proteins by a Specific Chemical Labeling.
Biochemistry Methods
Nanoparticles
Structural Biology
Journal
iScience
ISSN: 2589-0042
Titre abrégé: iScience
Pays: United States
ID NLM: 101724038
Informations de publication
Date de publication:
20 Dec 2019
20 Dec 2019
Historique:
received:
12
04
2019
revised:
26
10
2019
accepted:
12
11
2019
pubmed:
2
12
2019
medline:
2
12
2019
entrez:
2
12
2019
Statut:
ppublish
Résumé
Electron microscopy (EM) is a technology that enables visualization of single proteins at a nanometer resolution. However, current protein analysis by EM mainly relies on immunolabeling with gold-particle-conjugated antibodies, which is compromised by large size of antibody, precluding precise detection of protein location in biological samples. Here, we develop a specific chemical labeling method for EM detection of proteins at single-molecular level. Rational design of α-helical peptide tag and probe structure provided a complementary reaction pair that enabled specific cysteine conjugation of the tag. The developed chemical labeling with gold-nanoparticle-conjugated probe showed significantly higher labeling efficiency and detectability of high-density clusters of tag-fused G protein-coupled receptors in freeze-fracture replicas compared with immunogold labeling. Furthermore, in ultrathin sections, the spatial resolution of the chemical labeling was significantly higher than that of antibody-mediated labeling. These results demonstrate substantial advantages of the chemical labeling approach for single protein visualization by EM.
Identifiants
pubmed: 31786521
pii: S2589-0042(19)30475-4
doi: 10.1016/j.isci.2019.11.025
pmc: PMC6906691
pii:
doi:
Types de publication
Journal Article
Langues
eng
Pagination
256-268Informations de copyright
Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.
Références
Nat Chem Biol. 2014 Jul;10(7):512-23
pubmed: 24937069
Nat Chem Biol. 2015 Dec;11(12):917-23
pubmed: 26575238
Curr Opin Neurobiol. 2007 Jun;17(3):387-93
pubmed: 17499496
J Histochem Cytochem. 2000 Apr;48(4):471-80
pubmed: 10727288
Immunochemistry. 1971 Nov;8(11):1081-3
pubmed: 4110101
J Med Chem. 2014 Dec 11;57(23):10072-9
pubmed: 25375838
Histochem Cell Biol. 2018 Jan;149(1):3-13
pubmed: 29134300
Proc Natl Acad Sci U S A. 2017 Jun 27;114(26):E5246-E5255
pubmed: 28607047
Nat Chem. 2016 Feb;8(2):103-13
pubmed: 26791892
J Mol Biol. 2007 Sep 21;372(3):798-816
pubmed: 17681533
Bioconjug Chem. 2001 Nov-Dec;12(6):1051-6
pubmed: 11716699
Chem Commun (Camb). 2013 Jun 4;49(44):5022-4
pubmed: 23612601
Mol Biol Cell. 2007 Jun;18(6):2112-22
pubmed: 17392511
Cell. 2012 Aug 31;150(5):1068-81
pubmed: 22939629
Nat Commun. 2018 Aug 17;9(1):3307
pubmed: 30120230
Biochemistry. 2006 Apr 18;45(15):4785-96
pubmed: 16605247
Biochemistry. 1998 Jun 23;37(25):8965-72
pubmed: 9636038
J Am Chem Soc. 2010 Jul 14;132(27):9301-9
pubmed: 20568758
Nat Biotechnol. 2012 Nov;30(11):1143-8
pubmed: 23086203
Nat Chem. 2016 Feb;8(2):120-8
pubmed: 26791894
PLoS Biol. 2011 Apr;9(4):e1001041
pubmed: 21483721
J Cell Sci. 1995 Nov;108 ( Pt 11):3443-9
pubmed: 8586656
Mol Biosyst. 2016 May 24;12(6):1731-45
pubmed: 26960991
J Mol Biol. 1995 Nov 10;253(5):799-812
pubmed: 7473753
Inorg Chem. 2014 Feb 17;53(4):1816-23
pubmed: 24131471
J Med Chem. 2003 Jan 2;46(1):49-63
pubmed: 12502359
Curr Opin Chem Biol. 2010 Aug;14(4):475-80
pubmed: 20609616
J Am Chem Soc. 2016 May 4;138(17):5499-502
pubmed: 27082895
Science. 1998 Jul 10;281(5374):269-72
pubmed: 9657724
Curr Opin Biotechnol. 2017 Dec;48:61-68
pubmed: 28395178
FEBS Lett. 2015 Sep 14;589(19 Pt A):2590-602
pubmed: 25913176