Using cryo-EM maps to determine protein

transmembrane regions

Georgina Csizmadia

GPU Day 2019

memblob.hegelab.org

TheMemBlob database and server

Transmembrane proteins are significantdrug targets

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Membrane(bilayer)

Extracellularside

Intracellularside

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CFTR (PDBID: 5UAK)

Experimental data on the bilayer boundaries is sparse

CFTR (PDBID: 5UAK)

In silico methods are predictions

PPM MemProtMDTMDET

pdbtm.enzim.hu opm.phar.umich.edu memprotmd.bioch.ox.ac.uk

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Membrane embedment data is a blob in the EMD maps

CFTR (PDBID: 5UAK, EMD-8516)

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EMD

subtractionChimera

We extract theMemBlob from theEMD map

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PDB

CFTR (PDBID: 5UAK)

DMtheoretical

VMD

TMDET-PPM MEMPROTMD-PPM CG-PPM

In silico methods have similar resultsP

DB

ID

Csizmadia,G. et al. Computational and Structural Biotechnology Journal (2018)

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z, Å

α, deg

We convert the membrane blob to membrane boundaries

∑d(z, α, ∑d)

z

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water phaseUNK

hydrophobic core

interface region

Our pipeline is available as a webserver on memblob.hegelab.org

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Some EMD maps lack blobs

EMD-7338

EMD-7073

memblob

ABCC8/Kir6.2 complex

58

20 14

Database

Successfulrun

No visiblememblob

High noise& other

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a

Some of the experimental membrane blobs are thicker than expected

Membrane boundaries

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bMemBlob: solid lines

TMDET: dashed lines

c

Distribution of bilayer centres

TMDETz=0

d

Distribution of bilayerThickness

TMDET

MemBlob

Some proteins sit deep in the lipids

Lipid densitiesTMDET membrane boundaries

ABCC8/Kir6.2 (PDBID: 6BAA, EMD-7073)

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Does the membrane thickness depend on the type of membrane mimetics?

0

5

10

15

20

25

30

35

40

amphipol nanodisc micelle

Nu

mb

er o

f st

ruct

ure

s

similar membrane thickness

thicker more than 8 Å

thinner more than 8 Å

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• EM density maps contain extra information

•MemBlob is a pipeline to extract transmembrane

regions from EMD maps

• This provides high resolution experimental data on

transmembrane regions

Conclusions

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NKFIH-111678, NKFI-127961

CFF HEGEDU18I0, KIFÜ HPC

MTA Wigner GPU Laboratory

Semmelweis Science and Innovation Fund

FIKP 2018

Acknowledgments

memblob.hegelab.org

Eszter Katona

Gábor E. Tusnády

Tamás Hegedűs