Cryo-EM in the study of challenging systems: the human transcription pre-initiation complex.
Single particle cryo-Electron Microscopy (cryo-EM) is a technique that allows the structural characterization of macromolecules without the need for crystallization. For certain type of samples that are ideally suited for cryo-EM studies it has been possible to reach high-resolution structures following relatively standard procedures. Other biological systems remain highly challenging, even for cryo-EM. Challenges may involve the scarcity of the sample, poor stability of the complexes, and most often, the intrinsic flexibility of biological molecules. Among these challenging samples are large eukaryotic transcription complexes, which suffer from all such shortcomings. Here we report how we have recently tried to overcome those challenges in order to improve our structural understanding of the human transcription pre-initiation complex assembly and the transcription initiation process. Parallel efforts have also been carried out for budding yeast transcription initiation complexes, allowing comparisons that establish both the overall conservation and the specific structural differences between the two systems.
Molecular and Cell Biology Department and QB3 Institute, UC Berkeley, CA, USA; Howard Hughes Medical Institute, UC Berkeley, CA, USA; Molecular Biophysics and Integrative Bio-Imaging Division, Lawrence Berkeley National Lab, CA, USA. Electronic address: enogales@lbl.gov.
Eva
E
Nogales
Biophysics Graduate Group, UC Berkeley, CA, USA.
Robert K
RK
Louder
Department of Molecular Biosciences, Northwestern University, IL, USA.
Yuan
Y
He
D020285:Cryoelectron Microscopy / Q000379:methods*; D006801:Humans; D046911:Macromolecular Substances / Q000737:chemistry / Q000378:metabolism*; D061785:Transcription Initiation, Genetic
D046911:Macromolecular Substances
Nature. 2013 Jan 31;493(7434):699-702
23292512
Biochimie. 2013 Apr;95(4):680-91
23146842
Nature. 2016 Mar 31;531(7596):604-9
27007846
Cell. 2013 Jan 17;152(1-2):120-31
23332750
J Struct Biol. 2006 May;154(2):168-83
16510296
J Struct Biol. 2012 Dec;180(3):519-30
23000701
Nature. 2016 May 11;533(7603):353-8
27193681
Cell Res. 2014 Dec;24(12):1433-44
25412659
Nat Methods. 2013 Jun;10(6):584-90
23644547
Genes Dev. 2009 Jul 1;23(13):1510-21
19571180
Structure. 2008 May;16(5):673-83
18462672
Proc Natl Acad Sci U S A. 2015 Nov 3;112(44):13543-8
26483468
Structure. 2012 Nov 7;20(11):1823-8
23022349
J Struct Biol. 2006 Mar;153(3):284-99
16431136
Science. 1999 Dec 10;286(5447):2153-6
10591646
Science. 1999 Dec 10;286(5447):2151-3
10591645
Nature. 2013 Jan 17;493(7432):437-40
23151482
Structure. 2006 Mar;14(3):511-20
16531235
Transcription. 2013 May-Jun;4(3):120-6
23863784
Mol Cell. 2015 May 21;58(4):677-89
26000851
Nature. 2010 Jun 17;465(7300):956-60
20559389
J Struct Biol. 2013 Nov;184(2):321-8
24075951
Acta Crystallogr D Biol Crystallogr. 2010 Feb;66(Pt 2):213-21
20124702
Mol Cell. 2008 Jan 18;29(1):81-91
18206971
J Struct Biol. 2013 Sep;183(3):377-88
23872434
Cell. 1989 Feb 24;56(4):549-61
2917366
Nat Methods. 2007 Jan;4(1):27-9
17179934
J Struct Biol. 2006 May;154(2):184-94
16520062
Nat Methods. 2016 Jan;13(1):24-7
27110629
Proc Natl Acad Sci U S A. 2011 May 10;108(19):7745-50
21508329
Nature. 2016 May 11;533(7603):359-65
27193682
Nat Methods. 2006 Nov;3(11):917-22
17124735
Methods Enzymol. 2010;482:237-62
20888964
Methods. 2016 May 1;100:61-7
26884261
J Struct Biol. 2012 Dec;180(3):420-7
22902564
J Biol Chem. 2012 Aug 10;287(33):27580-92
22696218
Nature. 2013 Mar 28;495(7442):481-6
23446344