SASDAW5: XMRV RT + DNA/RNA hybrid (apo XMRV RT, XM + RNA_DNA hybrid substr...

Basic information

• Entry: Database: SASBDB / ID: SASDAW5

Sample

XMRV RT + DNA/RNA hybrid

• apo XMRV RT (protein), XM, Escherichia coli
• RNA_DNA hybrid substrate (other), PPT18

• Biological species: Escherichia coli (E. coli)
• Citation: Journal: Nucleic Acids Res / Year: 2013; Title: Structural analysis of monomeric retroviral reverse transcriptase in complex with an RNA/DNA hybrid.; Authors: Elzbieta Nowak / Wojciech Potrzebowski / Petr V Konarev / Jason W Rausch / Marion K Bona / Dmitri I Svergun / Janusz M Bujnicki / Stuart F J Le Grice / Marcin Nowotny / ; Abstract: A key step in proliferation of retroviruses is the conversion of their RNA genome to double-stranded DNA, a process catalysed by multifunctional reverse transcriptases (RTs). Dimeric and monomeric RTs have been described, the latter exemplified by the enzyme of Moloney murine leukaemia virus. However, structural information is lacking that describes the substrate binding mechanism for a monomeric RT. We report here the first crystal structure of a complex between an RNA/DNA hybrid substrate and polymerase-connection fragment of the single-subunit RT from xenotropic murine leukaemia virus-related virus, a close relative of Moloney murine leukaemia virus. A comparison with p66/p51 human immunodeficiency virus-1 RT shows that substrate binding around the polymerase active site is conserved but differs in the thumb and connection subdomains. Small-angle X-ray scattering was used to model full-length xenotropic murine leukaemia virus-related virus RT, demonstrating that its mobile RNase H domain becomes ordered in the presence of a substrate-a key difference between monomeric and dimeric RTs.

Contact author

• Petr Konarev (EMBL-Hamburg, European Molecular Biology Laboratory (EMBL) - Hamburg outstation, Notkestraße 85, Geb. 25A, 22607 Hamburg, Deutschland, Germany)

Models

• Model #142: ; Type: atomic / Software: CRYSOL / Chi-square value: 0.855625; Search similar-shape structures of this assembly by Omokage search (details)

Sample

• Sample: Name: XMRV RT + DNA/RNA hybrid / Sample MW: 90 kDa / Entity id: 103 / 112
• Buffer: Name: 10mM HEPES / pH: 6.5 / Composition: KCl 100.000 mM, glycerol 5.000 %

Entity #103

Name: XM / Type: protein / Description: apo XMRV RT / Formula weight: 75 / Num. of mol.: 1 / Source: Escherichia coli
Sequence:

MSHHHHHHSA TLNIEDEYRL HETSKEPDVP LGSTWLSDFP QAWAETGGMG LAVRQAPLII PLKATSTPVS IKQYPMSQEA RLGIKPHIQR LLDQGILVPC QSPWNTPLLP VKKPGTNDYR PVQDLREVNK RVEDIHPTVP NPYNLLSGLP PSHQWYTVLD LKDAFFCLRL HPTSQPLFAF EWRDPEMGIS GQLTWTRLPQ GFKNSPTLFD EALHRDLADF RIQHPDLILL QYVDDLLLAA TSEQDCQRGT RALLQTLGNL GYRASAKKAQ ICQKQVKYLG YLLKEGQRWL TEARKETVMG QPTPKTPRQL REFLGTAGFC RLWIPGFAEM AAPLYPLTKT GTLFNWGPDQ QKAYQEIKQA LLTAPALGLP DLTKPFELFV DEKQGYAKGV LTQKLGPWRR PVAYLSKKLD PVAAGWPPCL RMVAAIAVLT KDAGKLTMGQ PLVILAPHAV EALVKQPPDR WLSNARMTHY QAMLLDTDRV QFGPVVALNP ATLLPLPEKE APHDCLEILA ETHGTRPDLT DQPIPDADYT WYTDGSSFLQ EGQRRAGAAV TTETEVIWAR ALPAGTSAQR AELIALTQAL KMAEGKKLNV YTDSRYAFAT AHVHGEIYRR RGLLTSEGRE IKNKNEILAL LKALFLPKRL SIIHCPGHQK GNSAEARGNR MADQAAREAA MKAVLETSTL L

Entity #112

Name: PPT18 / Type: other / Description: RNA_DNA hybrid substrate / Formula weight: 15 / Num. of mol.: 1
Sequence:

5UAGUCUCCA GAAAAAGGGG GGAAUG35AT TCCCCCCTTT TTCTGGAGAC 3

Experimental information

• Beam: Instrument name: DORIS III X33 / City: Hamburg / 国: Germany / Shape: 0.6 / Type of source: X-ray synchrotron / Wavelength: 0.15 Å / Dist. spec. to detc.: 2.7 mm
• Detector: Name: Pilatus 1M-W / Pixsize x: 0.172 mm

Scan

Title: XM_DNA / Measurement date: Dec 8, 2011 / Storage temperature: 10 °C / Cell temperature: 10 °C / Exposure time: 15 sec. / Number of frames: 8 / Unit: 1/nm /

	Min	Max
Q	0.0855	6.032

Distance distribution function P(R)

Sofotware P(R): GNOM 4.5a / Number of points: 497 /

	Min	Max
Q	0.1718	2.893
P(R) point	32	528
R	0	11.57

Result

Type of curve: single_conc /

	Experimental	Porod
MW	90.5 kDa	-
Volume	-	155 nm3

	P(R)	Guinier
Forward scattering, I0	77	78.3
Radius of gyration, Rg	3.55 nm	3.5 nm

	Min	Max
D	-	11.5
Guinier point	8	99