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Department of Analytical Biochemistry

Webpage

www.zba.wbbib.uj.edu.pl

Head

Prof. Andrzej Kozik
room: 4.0.25 (A104), phone: +48 12 664 65 25

Staff

prof. dr hab. Adam Dubin, profesor emeritus
room: 4.0.21 (A118), phone: +48 12 664 65 11, e-mail: adam.dubin@uj.edu.pl

dr hab. Paweł Mak 
room: 4.0.22 (A119), phone: +48 12 664 65 38, e-mail: pawel.mak@uj.edu.pl

dr hab. Ibeth Guevara-Lora 
room: 4.0.26 (A103), phone: +48 12 664 65 27, e-mail: ibeth.guevara-lora@uj.edu.pl

dr hab. Benedykt Władyka 
room: 4.0.21 (A118), phone: +48 12 664 65 11, e-mail: benedykt.wladyka@uj.edu.pl

dr inż. Emilia Bonar 
room: 4.0.21 (A118), phone: +48 12 664 65 11, e-mail: emilia.bonar@uj.edu.pl

dr Michał Bukowski
room: 4.0.8 (A125), phone: +48 12 664 65 44, e-mail: m.bukowski@uj.edu.pl

dr Dorota Satała 
room: 4. 0.27 (A101), phone: +48 12 664 65 24, e-mail: dorota.satala@uj.edu.pl

dr inż. Michał Banasik
room: 4.0.7 (A124), phone: +48 12 664 65 06, e-mail: michal.banasik@uj.edu.pl

mgr Kamila Kulig 
room: 4. 0.8 (A125), phone: +48 12 664 65 44, e-mail: kamila.kulig@uj.edu.pl

dr Paulina Worsztynowicz
room: 4. 0.27 (A101), phone: +48 12 664 65 24, e-mail: paulina.worsztynowicz@uj.edu.pl

mgr Ewelina Wronowska 
room: 4. 0.8 (A125), phone: +48 12 664 65 44, e-mail: ewelina.wronowska@uj.edu.pl

PhD students

Marcin Hydzik, room: 4.0.8 (A125), phone: +48: 12 664 65 44
Monika Janczak, room: 4.0.7 (A124), phone: +48: 12 664 65 06  
Anna Mądry, room: 4.0.8 (A125), phone: +48: 12 664 65 43  
Kinga Chlebicka, room: 4.0.8 (A125), phone: +48: 12 664 65 43  
Aleksandra Żelazna, room: 4.0.8 (A125), phone: +48: 12 664 65 44  
Justyna Śmiałek, room 3.0.22 (B118), phone: +48: 12 664 63 41

Research topics

  • Molecular mechanisms of the host-pathogen interactions in bacterial and fungal infections
  • Biochemical and structural characterization of the virulence factors (mainly secreted proteases and cell-surface adhesins) of pathogenic bacteria (e.g. Staphylococci) and fungi (e.g. Candida yeasts)
  • Biochemical and structural characterization of proteinase inhibitors
  • Biochemical and structural characterization of antimicrobial peptides
  • Structural and functional modifications of proteins during inflammation
  • Functionality of the kinin-generating and blood-coagulation systems during inflammation and infections
  • Biochemistry of vitamins and coenzymes
  • Bacterial toxin-antitoxin systems

Methods and specialistic equipment

  • A broad spectrum of liquid chromatography methods. The Department is equipped with several low-pressure chromatographs (Pharmacia, BioRad), medium-pressure FPLC systems (Pharmacia, Knauer) and high-performance liquid chromatographs (Dionex, Shimadzu, Waters) with spectrophotometric and fluorimetric detectors.
  • Sequencing of proteins and peptides: The Department is equipped with an automatic protein sequencer Procise 491 (Applied Biosystems).
  • Mass spectrometry: The Department is equipped with a HCT Ultra ETDII (Bruker) mass spectrometer with ESI ion source and ion trap analyzer. This spectrometer is coupled with an ultrahigh-performance chromatograph Ultimate 3000 (Dionex). This LC-MS system allows to identify a variety of biologically active compounds by precise measurements of their molecular masses - directly or after enzymatic fragmentation. In our laboratories the spectrometer is applied for proteomic studies and for the identification and characterization of biologically active peptides.
  • Electrophoretic techniques. Proteins and peptides are analyzed using one- and two-dimension polyacrylamide gels (1D and 2D), both in mini and big formats. The comparative 2D electrophoresis is applied in studies concerning changes in intracellular and secretory proteomes of microorganisms, treated with different environmental factors. Selected proteins and peptides are then identified by direct automatic sequencing or mass spectrometry.
  • Molecular biology. We routinely perform nucleic acid amplification, molecular cloning, qualitative and quantitative determination of gene expression, construction of plasmid vectors, gene manipulations (introduction and knock-out). Additionally, we produce recombinant proteins in prokaryotic (Escherichia coli, Bacillus subtilis, Staphylococcus spp.) and eukaryotic (Pichia pastoris) hosts.
  • Dynamic light scattering (DLS). The Department is equipped with a DLS spectrometer (model DynaPro, Protein Solutions). This equipment allows to estimate the hydrodynamic radius of molecules in solutions. This parameter can then be used to calculate the molecular mass of the molecule, to determine homogeneity of the analyzed compounds as well as to trace changes in the size of molecules during aggregation or ligand binding.

Current projects

  1. Paweł Mak: Studies on biosynthesis regulation and on proinflammatory properties of isoforms of the peptide bacteriocin BacSp222. (2019-2022). Opus 16, National Science Centre (NCN).
  2. Emilia Bonar: Transcriptomic and proteomic analysis of the effects of mutations resulting in overproduction of PVL toxin in Staphylococcus aureus. (2019-2020). Miniatura 2, National Science Centre (NCN). 
  3. Benedykt Władyka: Studies of the role of toxin-antitoxin systems in antibiotic resistance in staphylococci.(2018-2021). Opus 13, National Science Centre (NCN).
  4. Andrzej Kozik: Cytoplasmic enzymes of pathogenic fungi Candida spp. that „moonlight” as adhesins on the cell wall – structural determinants of the novel function. (2017-2020). Opus 12, National Science Centre (NCN).
  5. Michał Bukowski: An investigation into novel mechanisms of regulation of gene expression in the context of interactions between host and Staphylococcus aureus cells during pathogenesis. (2016-2019). Sonata 11, National Science Centre (NCN).

Selected publications

  1. Mądry A, Jendroszek A, Dubin G, Wladyka B. (2019) Production of Lysostaphin by Nonproprietary Method Utilizing a Promoter from Toxin-Antitoxin System. Mol Biotechnol 61:774-782. 
  2. Nowakowski M, Jaremko Ł, Wladyka B, Dubin G, Ejchart A, Mak P. (2018) Spatial attributes of the four-helix bundle group of bacteriocins - The high-resolution structure of BacSp222 in solution. Int. J. Biol. Macromol. 107B, 2715-2724.
  3. Bonar E, Wojcik I, Jankowska U, Kedracka-Krok S, Bukowski M, Polakowska K, Lis MW, Kosecka-Strojek M, Sabat AJ, Dubin G, Friedrich AW, Miedzobrodzki J, Dubin A, Wladyka B (2016) Identification of secreted exoproteome fingerprints of highly-virulent and non-virulent Staphylococcus aureus strains. Front Cell Infect Microbiol 6:51.
  4. Wladyka B, Piejko M, Bzowska M, Pieta P, Krzysik M, Mazurek Ł, Guevara-Lora I, Bukowski M, Sabat AJ, Friedrich AW, Bonar E, Międzobrodzki J, Dubin A, Mak P (2015) A peptide factor secreted by Staphylococcus pseudintermedius exhibits properties of both bacteriocins and virulence factors. Sci Rep 5:14569.
  5. Bochenska O, Rapala-Kozik M, Wolak N, Kamysz W, Grzywacz D, Aoki W, Ueda M, Kozik A (2015) Inactivation of human kininogen-derived antimicrobial peptides by secreted aspartic proteases produced by the pathogenic yeast Candida albicans. Biol Chem 396:1369-75.
  6. Kozik A, Gogol M, Bochenska O, Karkowska-Kuleta J, Wolak N, Kamysz W, Aoki W, Ueda M, Faussner A, Rapala-Kozik M (2015) Kinin release from human kininogen by 10 aspartic proteases produced by pathogenic yeast Candida albicans. BMC Microbiol 15:60.
  7. Kozik A, Karkowska-Kuleta J, Zajac D, Bochenska O, Kedracka-Krok S, Jankowska U, Rapala-Kozik M (2015) Fibronectin-, vitronectin- and laminin-binding proteins at the cell walls of Candida parapsilosis and Candida tropicalis pathogenic yeasts. BMC Microbiol 15:197.
  8. Karkowska-Kuleta J, Kozik A. (2014) Moonlighting proteins as virulence factors of pathogenic fungi, parasitic protozoa and multicellular parasites. Mol Oral Microbiol 29:270-283.
  9. Guevara-Lora I, Stalinska K, Augustynek B, Labedz-Maslowska A. (2014) Influence of kinin peptides on monocyte-endothelial cell adhesion. J Cell Biochem 115:1985-1995.
  10. Bukowski M, Lyzen R, Helbin WM, Bonar E, Szalewska-Palasz A, Wegrzyn G, Dubin G, Dubin A, Wladyka B. (2013) A regulatory role for Staphylococcus aureus toxin-antitoxin system PemIKSa. Nat Commun 4:2012.
     

Batchelor/master thesis topics

  • Isolation and biochemical characterization of proteolytic enzymes
  • Protein expression in heterological bacterial expression systems
  • Cloning, expression and purification of recombinant proteins
  • Antibacterial peptides: isolation, purification, sequencing and biochemical characterization
  • Bacteriocins: isolation, purification, sequencing and biochemical characterization
  • New techniques in protein and peptide chemistry
  • Interaction of the contact system proteins – kininogen, prekallikrein and factor XII – with the surface of selected Candida spp.
  • Kinin generation by secreted proteinases from Candida spp.
  • Mechanism of kinin generation in various cell models
  • Expression of genes and proteins involved in the regulation of inflammation, especially in relation to the kinin generation system
  • Toxin-antitoxin systems: identification, characteristics, application in biotechnology

  • Regulators of gene expression in bacteria

Requirements for candidates

Interests in theoretical aspects and in experimental work concerning biochemistry, genetic engineering and physicochemistry.