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PD Dr. Basem Soboh

Microbiology, Biochemistry and Genetic Biophysics

Research Focus
In vitro
biosynthesis of complex Fe-S cofactors

My name is Basem Soboh. The major focus of my current work is to understand how complex cofactors of gas processing metalloproteins are assembled. I was born in Gaza-Palestine, where I completed my Bachelor degree of Microbiology & Chemistry, then I completed my diploma and doctoral studies at the MPI-Marburg in the group of Rolf Thauer. I studied a wide combination of subjects including Microbiology, Biochemistry, Genetics and Virology. During my post-doc at UC-Berkeley I investigated the in vitro biosynthesis of the FeMo cofactor of Nitrogenase. During my Habilitation at Martin Luther University, an in vitro Synthesis of active [NiFe]-hydrogenase has been developed. Since 2016 I started as a group leader in the laboratories of Heberle for Experimental Biophysics and Schlesinger for Genetic Biophysics. Using a combinations of molecular biology, anaerobic protein biochemistry and biophysical methods, we are currently investigating the biogenesis of [NiFe]-cofactor. Further details can be found in my CV.


Fachbereich Physik

Institut für experimentelle Physik

Experimentelle Molekulare Biophysik

Wissenschaftlicher Mitarbeiter

Arnimallee 14
Raum -1.2.06
14195 Berlin
(030) 838-453337


2016–present: Freie Universität Berlin
Group leader at the Department of Physics, Genetic Biophysics
Topic: In vitro biosynthesis of [NiFe]-hydrogenases

2008–2015: Martin Luther University
Group leader at the Department of Biology, Institute of Microbiology
Topic: In vitro biosynthesis of complex Fe-S cofactors

2005–2008: University of California, Berkeley
Postdoctoral Scholar at the Department of Plant and Microbial Biology (with Prof. Paul W. Ludden)
Topic: Biosynthesis of the iron-molybdenum cofactor of nitrogenase

2004–2005: MPI for Terrestrial Microbiology
Postdoctoral Scholar, Department of Biochemistry (with Prof. Rolf Thauer)
Topic: Characterization of energy-conserving [NiFe]-hydrogenases and CO-dehydrogenases



Habilitation in Microbiology 2016, Martin Luther University (Institute of Biology-Microbiology)

Ph.D. (Dr. rer. nat.) 2004, Department of Biochemistry at the MPI Marburg (Thauer Lab)

Diploma in Biology 2001, Philipps-University Marburg (Microbiology, Biochemistry, Genetics & Virology)

Bachelor of Science 1996, Alazhr University-Gaza (Microbiology & Chemistry)

Understanding microbial metabolism and energy conversion with respect to gas-processing enzymes e.g. energy converting membrane bound [NiFe] hydrogenases (H2), NADH-dependent Fe-hydrogenase, CO-dehydrogenase (CO), formate-dehydrogenase (CO2) and nitrogenase (N2). Elucidating the biosynthesis and catalytic mechanism of these enzymes at the molecular level (bio-catalysis).

A major focus of my current work is understanding how the cofactor of [NiFe]-hydrogenases is assembled. Over the last few years, we have developed a novel in vitro maturation system for the synthesis of active [NiFe]-hydrogenase using only purified components. Our in vitro reconstitution system provides the possibility to study the maturation steps individually and to observe the stepwise synthesis and assembly of the hydrogenase cofactor in real time. The in vitro system will be important for elucidating the biogenesis of [NiFe]-cofactor at the molecular level.

Our biochemical-genetic strategy involves isolation of the maturation proteins, then following the stepwise biosynthesis and assembly of cofactors in real time. We are using a broad range of methodologies. This includes manipulation of genes, overexpression and the anaerobic purification of maturation protein complexes that required for the in vitro reconstitution of the pathway for cofactor biosynthesis. The analytical methods include anoxic enzyme kinetics, FPLC, metabolite analysis (HPLC, GC), functional protein-protein interaction (thermophoresis), metal detection (ICP-MS), and native gel electrophoresis. Spectroscopic methods include UV/Vis-, electron paramagnetic resonance (EPR)-, Mössbauer-, resonance Raman-, and Fourier-transform infrared (FTIR) spectroscopy. Furthermore, we apply anaerobic crystallization, electrophysiology experiments using planar lipid bilayers and protein film electrochemistry in order to record the catalytic currents of enzyme complexes. Large size membrane proteins and enzyme complexes will be determined using cryo electron microscopy.

Working model for maturation of the large subunit into functional [NiFe]-hydrogenase. (1) The HybG-HypD complex is formed upon contact with the [Fe]CO2 carrying HybG dimer. (2) The CO2 ligand may undergo an ATP-dependent reduction to CO catalyzed by HybG-HypD complex. (3) The maturation proteins HypF and HypE catalyze the synthesis and transfer of CN ligands to the CO-modified iron ion and formation of the [Fe](CN)2CO moiety on the HybG-HypD complex. (4) It is unclear how the [Fe](CN)2CO moiety is incorporated into the large subunit. (5) Nickel insertion by concerted activity of HypA, HypB, and SlyD. (6) Endoproteolytic cleavage of the C-terminal peptide, associated with conformational changes of the protein. (7) Dimerization of large and small subunit forms functional [NiFe]-hydrogenase.

Selected Publications

1- Soboh, B*, Adrian, L. and Stripp, S. T. (2022) An in vitro reconstitution system to monitor iron transfer to the active site during the maturation of [NiFe]-hydrogenase, Journal of Biological Chemistry, doi: https://doi.org/10.1016/j.jbc.2022.102291.

2- Stripp, S. T., Oltmanns, J., Muller, C. S., Ehrenberg, D., Schlesinger, R., Heberle, J., Adrian, L., Schunemann, V., Pierik, A. J., and Soboh, B*. (2021) Electron inventory of the iron-sulfur scaffold complex HypCD essential in [NiFe]-hydrogenase cofactor assembly. Biochemical journal 478, 3281-3295

3- Senger M., Laun K., Soboh B. and Stripp S.T. (2018) Infrared Characterization of the Periplasmatic O2-sensitive [NiFe]-hydrogenase from E. coli. Catalysts, (8), 530

4- Senger M., Stripp S.T. and Soboh B*. (2017) Proteolytic Cleavage Orchestrates Cofactor Insertion and Protein Assembly in [NiFe]-hydrogenase Biosynthesis. J Biol Chem. (28):11670-11681.

5- Stripp ST., Lindenstrauss U., Sawers RG. and Soboh B*. (2015) Identification of an Isothiocyanate on the HypEF Complex Suggests a Route for Efficient Cyanyl-Group Channeling during [NiFe]-Hydrogenase Cofactor Generation. PLoS One e0133118. doi: 10.1371/journal.pone.0133118.

6- Stripp ST., Lindenstrauss U., Granich C., Sawers RG. and Soboh B*. (2014) The influence of oxygen on [NiFe]-hydrogenase cofactor biosynthesis and how ligation of carbon monoxide precedes cyanation. Plos ONE 9 e107488. doi: 10.1371/journal.pone.0107488.

7- Soboh B*., Lindenstrauss U., Granich C., Javaid M., Herzberg M., Claudia T. and Stripp ST. (2014) [NiFe]-hydrogenase maturation in vitro: analysis of the roles of the HybG and HypD accessory proteins. Biochemical journal. 1;464(2):169-77.

8- Soboh B., Stripp ST., Bielak C., Lindenstrauß U., Braussemann M., Javaid M., Hallensleben M., Granich C., Herzberg M., Heberle J. and Sawers RG. (2013) The [NiFe]-hydrogenase accessory chaperones HypC and HybG of Escherichia coli are iron- and carbon dioxide-binding proteins. FEBS Lett. 19;587(16):2512-6.

9- Soboh B. and Sawers RG. (2013) [NiFe]-hydrogenase cofactor assembly. In: Encyclopedia of Inorganic and Bioorganic Chemistry - Metals in Cells, Chapter eibc2154 ISBN: 9781119951438. doi: 10.1002/9781119951438.eibc2154.

10- Stripp ST., Soboh B., Lindenstrauss U., Braussemann M., Herzberg M., Nies DH. , Sawers RG. and Heberle J. (2013) HypD is the Scaffold Protein for Fe-(CN)2CO Cofactor Assembly in [NiFe]-Hydrogenase Maturation. Biochemistry, 52 (19), 3289–32962

11- Trchounian K., Soboh B., Sawers RG. and Trchounian A. (2013) Contribution of hydrogenase 2 to stationary phase H2 production by Escherichia coli during fermentation of glycerol. Cell. Biochem. Biophys. 66-(1)103-108.

12- Soboh B., Stripp ST., Muhr E., Granich C., Braussemann M., Herzberg M., Heberle J. and Sawers RG. (2012) [NiFe]-hydrogenase maturation: isolation of a HypC-HypD complex carrying diatomic CO and CN- ligands.FEBS Lett., 586(21) 3882-3887

13- Soboh B., Kuhns M., Braussemann M., Waclawek M., Muhr E., Pierik AJ. and Sawers RG. (2012) Evidence for an oxygen-sensitive iron-sulfur cluster in an immature large subunit species of Escherichia coli [NiFe]-hydrogenase 2. Biochem Biophys Res Commun. 424(1),158-163

14- Petkun S., Shi R., Li Y., Asinas A., Munger C., Zhang L., Waclawek M., Soboh B., Sawers RG. and Cygler M. (2011) Structure of Hydrogenase Maturation Protein HypF with Reaction Intermediates Shows Two Active Sites. Structure 19 (12), 1773–1783

15- Pinske C., Krüger S., Soboh B., Ihling C., Kuhns M., Braussemann M., Jaroschinsky M., Sauer C., Sargent F., Sinz A. and Sawers RG. (2011) Efficient electron transfer from hydrogen to benzyl viologen by the [NiFe]-hydrogenases of Escherichia coli is dependent on the coexpression of the iron-sulphur cluster-containing small subunit. Arch. Microbiol.193(12),893-903

16- Soboh B., Pinske C., Kuhns M., Waclawek M., Ihling C., Trchounian K., Trchounian A., Sinz. A., and Sawers RG. (2011) The respiratory molybdo-selenoprotein formate dehydrogenases of Escherichia coli have hydrogen: benzyl viologen oxidoreductase activity. BMCMicrobiol.11:173

17- Soboh B., Krüger S., Kuhns M., Pinske C., Lehmann A. and Sawers RG. (2010) Development of a cell-free system reveals an oxygen-labile step in the maturation of [NiFe]-hydrogenase 2 of E. coli.FEBS Lett. 584 (18), 4109-4114

18- Soboh B., Boyd ES., Zhao D, Peters JW. and Rubio LM. ( 2010) Substrate specificity and evolutionary implications of a NifDK enzyme carrying NifB-co at its active site. FEBSLett. 584(8),1487-92

19- Rubio LM., Hernández JA., Soboh B., Zhao D., Igarashi RY. , Curatti L. and Ludden PW. (2008). The Role of Nif Proteins in Nitrogenase Maturation. Plant Science and Biotechnology in Agriculture, Book: Biological Nitrogen Fixation, Volume 42, pp 325-328

20- Curatti L., Hernandez JA., Igarashi RY., Soboh B., Zhao D. and Rubio LM. (2007). In vitro synthesis of the iron-molybdenum cofactor of nitrogenase from iron, sulfur, molybdenum and homocitrate using purified proteins. Proc. Natl. Acad. Sci. 104 (45), 17626-31

21- George SJ., Igarashi RY., Piamonteze C., Soboh B., Cramer SP. and Rubio LM. (2007) Identification of a Mo-Fe-S cluster on NifEN by Mo K-edge EXAFS. J. Am. Chem. Soc.,129(11),3060-3061

22- Hernández JA., Igarashi RY., Soboh B., Curatti L., Dean DR., Ludden PW. and Rubio, LM. (2006) NifX and NifEN exchange biosynthetic precursors of the iron-molybdenum cofactor of nitrogenase. Mol. Microbiol.,63 (1),177-92

23- Soboh B., Igarashi RY., Hernandez JA. and Rubio LM. (2006) Purification of a NifEN protein complex that contains bound Mo and a FeMo-co precursor from an Azotobacter vinelandii ΔnifHDK strain. J. Biol. Chem., 281, 36701-36709cofactor of nitrogenase. Mol. Microbiol.,63 (1),177-92

24- Soboh B., Forzi L., Stojanowic A. and Hedderich R. (2004) Energy-converting [NiFe] hydrogenases from archaea and bacteria: ancestors of complex I. Biochimica et Biophysica Acta (BBA) – Bioenergetics. 1658

25. Soboh B., Linder D. and Hedderich R. (2004) A multisubunit membrane-bound [NiFe] hydrogenase and a NADH-dependent Fe-only hydrogenase in the fermenting bacterium Thermoanaerobacter tengcongensis Microbiology 150, 2451-2463

26. Soboh B., Linder D. and Hedderich R. (2002) Purification and catalytic properties of a CO-oxidizing:H2 evolving enzyme complex from Carboxydothermus hydrogenoformans Eur.J. Biochem. 269, 5712-21.