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03/11/2010 1 What’s new in Structural Biology? Vernon Smith Bruker AXS

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Page 1: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20101

What’s new in Structural Biology?

Vernon SmithBruker AXS

Page 2: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20102

What’s new in Structural Biology at Bruker?

Vernon SmithBruker AXS

Page 3: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20103

Welcome to the Bio Users Meeting 2010

Thank you to our session chairs Frank von Delft, Roman Fedorov and Michael Groll

Thank you to our speakers

Thank you to our organisers Martin Adam and Anja Griessmeijer

Thank you for attending:

Czech Republic, Estonia, Finland, France, Germany, India, Ireland, Israel, Italy, Kuwait, The Netherlands, Poland, Portugal, Singapore, Slovakia, Slovenia, South Africa, Spain, Sweden, United Kingdom, USA,

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03/11/20104

Users Meetings; continuing …

University of Alabama, AL S Ealick, C Smith, E Westbrook Feb. 26-29, 1989 CARB, MD G Gilliland,A Howard,G Cohen,L Hannick,A Wlodawer,M Amzel Feb. 4-6, 1990 Scripps, CA I Wilson, E Stura, D McRee, M Pique May 5-7, 1991 Purdue University, IN Wladek Minor, Ivan Rayment May 3-5, 1992 GBF Braunschweig, Germany D Schomburg, H.J.Hecht, B.Hofmann Apr. 28-30, 1993 University of Pittsburgh,PA John Rose, William Furey Apr. 28-30, 1994 University of Georgia, Athens, GA B-C Wang, John Rose Apr. 17-19, 1997 Scripps, La Jolla, CA Duncan McRee, Dave Stout Feb. 21-23, 1998 University of Bergen, Norway (ESUM98) Karl Tornroos Sep.11-13, 1998 University of Toledo, OH Alan Pinkerton June 9-11, 1999 Texas A&M University, TX Joe Reibenspies Mar. 16-17, 2000 University of Wisconsin/Bruker, WI Ilia Guzei, Charles Campana May 20-22, 2001 University of Wisconsin/Bruker, WI Ilia Guzei, Charles Campana May 18-20, 2003 University of Wisconsin/Bruker, WI Ilia Guzei Sept 20-22, 2005 Bruker AXS Delft Frank van Meurs Sept 18-19, 2006 University of Wisconsin/Bruker, WI Ilia Guzei Sept 16-18, 2007 Universität Göttingen George Sheldrick Oct 10-13, 2007 Bruker AXS Delft Frank van Meurs Sept 29-1, 2008 University of Wisconsin/Bruker, WI Ilia Guzei June 1-2, 2009 Bruker AXS Karlsruhe Martin Adam Sept 20-23, 2010

Page 5: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20105

Bruker News

Single crystal has moved to Bruker AXS Headquarters, Karlsruhe

New Bio Application Scientist – Dr Marianna Biadene

Strengthening service & support…

Page 6: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20106

Bruker AXS, Karlsruhe, Germany

Business Manager Eric Hovestreydt

Product Manager, Marketing Martin Adam

Business Development Vernon Smith

(Biological Crystallography) Bernd Hinrichsen (X2S)

Martin Adam

Vernon Smith

Eric Hovestreydt

Bernd Hinrichsen

Page 7: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20107

Service Karl-Heinz Bast Oliver Knaus Markus Nuesse Klaus Orth Felix Schmidt Daniel Stern

Bruker AXS, Karlsruhe, Germany

Karl-Heinz Bast Oliver Knaus Markus Nüsse

Klaus Orth Felix Schmidt Daniel Stern

Page 8: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20108

Bruker AXS Delft, the Netherlands

R&D Jaap Toorn

Application Leo Straver

Service Cees Baas Henk Lief Dirk Parlevliet Frank Vredenbregt

Cees Baas Henk Lief

Dirk ParlevlietFrank Vredenbregt

Leo Straver

Jaap Toorn

Page 9: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/20109

IMCB, Singapore Manchester Interdisciplinary

Biocentre, UK University of Vienna, Austria

Protein Crystallography workshops

“… the workshop was a success and appreciated by all involved.”

(Bob Robinson, IMCB, Singapore)

Page 10: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201010

Product News

MX OPTICS

NEW SOURCES

Page 11: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

X8 PROSPECTOR

IS with Quazar MX optics• No external cooling• No 3-phase power• No filament changes

Apex II detector

• High spatial resolution• Highest sensitivity• Lowest noise• Fast readout

Kappa Goniometer

• Easy sample handling• Powerful data collection tool

Page 12: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

Source Comparison: IS

Page 13: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201013

• Small thaumatin crystal: 105 83 50 µm3 , = 0.5°, 120 s/°, D= 120

Resolution 2.05 Å<I/> = 12.2 (2.7)Rmerge = 0.097 (0.45)

Quazar

Resolution 1.92 Å<I/> = 13.2 (2.9)Rmerge = 0.078 (0.41)

Quazar MX

Incoatec Multilayer Optics: Comparison

Page 14: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

X8 PROSPECTORLonger Axis at High Divergence

• Wild-Type dTDP-4-keto-6-deoxy-D-glucose-3,4-ketoisomerase in complex with TDP

• Source Aneurinibacillusthermoaerophilus

• Beam divergence 7 mrad• Space group P41212• Unit Cell: a=b=61.7, c=201.6 Å• Hazel Holden, UW-Madison

High spatial resolution of the APEX II CCD coupled with the smaller focus of the MX optic allows separation of a longer axis even with high divergence

Page 15: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

System Comparison: X8 PROSPECTOR

• Data were collected on the same frozen lysozyme and thaumatinsamples.

• The Rotating Anode Generator data were collected at a user site; the ImS data were collected at Bruker AXS Madison applications lab

• Match the resolution and redundancy then compare the statistics

Source RAG IS

Optics Multilayer optics

Quazar

Power load 5 kW 30 W

Detector Image plate APEXII CCD

Comparison of the IS with a conventional 5.4 kW rotating anode

Page 16: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

Comparison Results: Thaumatin

Thaumatin crystal 1 IS RAG

Degrees collected (0.5o per frame) 270 100

Collection time (hrs) 3 9

Exposure per frame (sec) 15 60

Resolution (Å) 2.04 2.04

Rsymm 0.052 (0.13) 0.060 (0.24)

I/sigma(I) 24.7 (9.3) 11.4 (7.4)

Completeness (%) 99.1 (98.3) 98.8 (99.9)

Redundancy 6.2 6.6

Thaumatin crystal 2 IS RAG

Degrees collected (0.5o per frame) 270 210

Collection time (hrs) 3 17

Exposure per frame (sec) 15 40

Resolution (Å) 1.85 1.85

Rsymm 0.077 (0.33) 0.080 (0.69)

I/sigma(I) 14.6 (4.3) 8.5 (2.3)

Completeness (%) 99.9 (99.9) 99.9 (100)

Redundancy 5.5 7.7

In each case the X8 PROSPECTOR produced better data

• Lower Rmerge

• Higher signal to noise

• Data were collected up to 5 times faster

Page 17: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

X8 PROSPECTOR Ligand binding studies

Resolution %Complete Redundancy Mean I/I Rmerge

Inf - 2.10 99.2 (99) 7.19 (4.9) 20.89 (3.2) 0.064 (0.40)

Data collection parameters

• Detector DX 100 mm

• Exposure time 40 sec

• Rotation angle 0.3°

• Degrees collected 540°

• Total time ~21 hrs

• Synchrotron data collected at SBC 19-BM to 1.5 Å

Page 18: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201018

X8 PROTEUM

High brilliance MICROSTAR rotating anode

Helios MX multi-layer optics

Platinum135 CCD detector

Kappa goniometer

Low Temp

Superb performance for

• High quality data

• Protein screening

• In-house SAD phasing

Page 19: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201019

NEW Turbo X-Ray Source

Compact microfocus rotating anode

Directly-mounted onto goniometer-base

= extremely stable

Simple maintenance

• Pre-aligned filaments

• Anode exchange

• Ferroseal exchange

HELIOS or HELIOS MX optics

Performance comparable with MICROSTAR

Page 20: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201020

Source Comparison: MICROSTAR MX

Intensity similar to 2nd generation synchrotron beamlines

Page 21: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201021

X8 PROTEUMHigh Resolution Data Collection

a=b=68.08 Å c=102.01 Å Source Tritirachium Crystal size 0.3x0.22x0.15 mm Exp time (sec/deg) 90 Rmerge (%) 4.1(20.4) II) 24.6(6.71) Completeness(%) 99.4(98.1) Redundancy 8.0(5.23) Collection time (hrs)40

Proteinase K at 0.95 Å

Red election density is a 2FomFc map contoured at 1, yellow density is an FomFc map at 3 showing possible hydrogen atom positions

Page 22: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201022

X8 PROTEUMHigh Resolution Data Collection

Page 23: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201023

X8 PROTEUMSulfur SAD Phasing

Source Saccharomyces cerevisiae

48 kD, 428 aa, monomer in ASU

Unit cell: a=b=85.6, c=132.8 Å,

Space group: P3121

Solvent content 56%

Substructure solved at 3.5 Å

11 Cys, 8 Met found 13 sulfurs

Redundancy ~ 50 fold to 2.5 Å

Data collected to 2.2 Å

SHELXE fit 85% of the backbone in ~30 minutes

VPS15 WD Repeat Domain

Dr. Janeen Van Hooke, Sondek lab, UNC

Page 24: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201024

X8 PROTEUMSulfur SAD Phasing

Structural genomics target PF1117

• Source Pyrococcus furiosus

• 85 aa, 3 met• Unit Cell: a=b=47.5, c=81.9 Å• Space group P41212

Resolution %Complete Redundancy Mean I/s R(int)

Inf ‐ 4.52 99.9 103 247.34 0.02154.52 ‐ 3.51 100 114.22 228.97 0.02733.51 ‐ 3.03 100 115.81 168.76 0.03773.03 ‐ 2.74 100 83.89 81.81 0.06232.74 ‐ 2.53 100 46.24 38.88 0.10492.53 ‐ 2.36 100 25.41 19.72 0.15882.36 ‐ 2.23 100 11.62 8.7 0.23362.23 ‐ 2.10 100 6.88 5.49 0.21882.10 ‐ 2.00 100 6.43 3.34 0.3214

Inf ‐ 2.00 100 53.56 82.84 0.0275

Data reduction statistics

B.C. Wang, University of Georgia

Page 25: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201025

X8 PROTEUMSulfur SAD Phasing

Substructure determination: SHELXD

•Data range 40 – 3.0 Å•3 peaks above background located

• Correlation coefficient ~50• MET23, MET68, 1 unknown

Phasing and backbone trace: SHELXE

•Phases extended to 2.0 Å•64 main chain residues found (75%)•Map generated from experimental phases contoured at 1σ•Picture shows β-strand region of molecule

Page 26: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201026

Sulfur SAD Phasing on X8 PROTEUMHetL, Pentapepide Repeat Protein

Source Anabaena sp.

237 resides: 4 CYS, 2 MET

Unit cell: a=68.2, b=93.7, c=101.3 Å, I222

Sub structure determined at 2.7 Åusing SHELXD

Phases were extended to 2.0 Åand the main chain atoms for all 237 residues were determined using a development version of SHELXE

Main chain fit in ~20 minutes

Structure from Mike Kennedy’s lab at Miami University of Ohio

Page 27: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201027

Sulfur SAD PhasingHetL, Pentapepide Repeat Protein

Page 28: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201028

X8 PROTEUMSAD Phasing of D-Glycerate 2-Kinase

Source E. coli 42 kD protein, 408 aa

Data resolution to 1.8 Å a=225.2,b=63.4,c=62.0 =95.6o

Crystals soaked with PtCl4, produced one heavy atom site

Overall Rsym 4.3% Average redundancy 8 fold Experimental map from one data

set collected in less than 24 hrs Chain traced by Arp/Warp overnight

Page 29: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201029

X8 PROTEUMUnit cell with Long Axis

P6122 a=b=70.44, c=439.6

Detector distance 200 mm Exposure time 450 sec/deg Rotation angle 0.2 Degrees collected 114 Time required ~15 hrs Divergence 3 mrad

Rmerge 6.95 (25.2) I/I 11.4 (3.2) Completeness 87% (71) Redundancy 3.0 (1.5)

Greg Petsko, Brandeis University

Page 30: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201030

Fast Screening30 second Lysozyme Dataset to 2.0 Å

Exposure time 0.5 seconds

Rotation width 1°Data collected 60°Total time 2.5 minutes

Total measurement time 30 seconds

Resolution range (Å) 22 - 2.00

Rmerge (%) 6.34 (17.89)

I/σ(I) 15.73 (4.24)

Completeness (%) 97.6 (85.1)

Redundancy 3.68 (1.73)

Page 31: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201031

Metal jet microfocus source

e-beam

liquid metalanode

x-rays

X-Ray source technology

Conventional microfocus source

solid metalanode

e-beam x-rays

Metal jet microfocus source

e-beam

liquid metalanode

x-rays

99.5 % of electron power becomes heat

Solid target close to melting

few 10 kW/mm2 define technology limit

high-speed liquid-metal-jet anode

Anode is self healing

> 500 kW/mm2

Metal jet microfocus source

e-beam

liquid metalanode

x-rays

Page 32: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201032

Shutter

Montel Mirror

e- optic

Metal-Jet X-ray Source (MJXS)Development model

Prototype installation, R & D Lab, Karlsruhe

Page 33: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201033

Target: Galinstan (eutectic mix of Ga(>75%), In, Sn) Generator Max. Energy: 60kV Kα line (Ga): 9.2keV

Development modelSource spectrum

Photon Energy

Inte

nsity

[arb

.U.]

Ga Kα 9,2 keV

Ga Kβ 10,3 keV

In Kα 24 keV

Sn Kα 25.3 keV

In Kβ 27.3 keV

Sn Kβ 28.5 keV

Page 34: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201034

66

1178

160

600

0 100 200 300 400 500 600 700

Intensity x 109 (X-rays/sec/mm2)

Classic 5 kW RAG/multilayer optics

IuS - Quazar

IuS - Quazar MX

EXCILLUM MJXS

MICROSTAR-H

MICROSTAR-MX

>5.0 x 1011

ph/mm2/s

X-ray SourcesComparison of flux through a 0.1 mm aperture

Beam FWHM @ sample, μm

300

240

110

260

160

~100

Page 35: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201035

Bruker AXS SourcesMICROSTAR

•Microfocus rotating anode generator

•Highest intensity RAG

•Dual-port

TURBO X-RAY SOURCE

•Microfocus rotating anode generator

•Compact, stable, ease-of-use

•Single-port

IS

•Microfocus sealed tube source

•Ultra reliable

•Flux greater than 5 kW RAG

METAL-JET X-RAY SOURCE

•10x brighter than current home sources

•Stability

•Low-maintenance

Page 36: What’s new in Structural Biology? - X-ray Homexray.chem.wisc.edu/Resources/Meetings/Bruker18/05_VRS_Bio-news.… · What’s new in Structural Biology? Vernon Smith Bruker AXS

03/11/201036

www.bruker-axs.com