Protein Engineering Laboratory

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Protein Engineering Laboratory

The Protein Engineering and Particle Reacting Laboratory is comprehensively equipped with the most modern equipment for testing properties and functions of proteins with biochemical and biophysical methods. We offer the possibility of recombinant proteins production and purification from different expression platforms. Moreover, the laboratory is prepared for the analysis of reactions occurring between proteins, proteins and DNA, ligands and peptides. We have at our disposal specialist equipment enabling measurement of bioactivity and kinetic parameters in a wide range from pM to mM.

We perform testing within scientific projects and commercial orders.

Production and purification of recombinant proteins from different expression systems

Qualitative and quantitative protein bioanalysis:

  • Identification (MS, MS/MS)
  • Purity (PAGE, WB, SEC, UV-Vis, HPLC/FPLC)
  • Activity (SPR, BLI, MST, ELISA, AlphaScreen, AlphaLISA, UV-NIR, FRET, TR-FRET, TRF, FP)
  • Aggregation/oligomerization (DLS, SEC)
  • Stability (DSC, CD)
  • Structural analysis (CD)

Protein designing/modification in order to obtain desired properties/activities

Testing of molecular reacting

size of analysed reagents from ions to viruses

Kinetics and reaction mechanism

Kd, kon, koff

Kd, DH, DS, stoichiometry

Enzyme kinetics parametrization

kcat, Vmax, Km, IC50, Ki

Bioactivity testing

SPR, BLI, MST

ELISA, AlphaScreen, AlphaLISA

UV-NIR, FRET, TR-FRET, TRF, FP

Selection, characterisation and optimization of potential bio therapeutics

Apparatus for measuring interaction between molecules:

Biacore T200

Apparatus using surface plasmon resonance (SPR)

 

Octet RED384

Device using layered interferometry (BLI)

 

 Monolith NT.115Pico

Device using microscale thermophoresis (MST)

 

Dionex Ultimate 3000

Dionex Ultimate 3000

 

AKTA Explorer

AKTA Explorer

 

Tecan FreedomEVO 150

Multifunctional workstation for cleaning biopreparations with a plate reader Infinite M1000 Pro

Tecan FreedomEVO 200

Automatic station for testing ligand receptor interactions with a multifunctional reader Infinite M200 Pro

 

MicroCal VP-Capillary DSC

Automated differential scanning calorimeter

 

MicroCal ITC200

Isothermal titration calorimeter

 

Zetasizer DLS/SLS Nano ZS

Zetasizer DLS/SLS Nano ZS

 

Jasco J-815

Apparatus for measuring the rotation of CD light

Plate readers:

PE Enspire with modul Corning Epic, PE Victor X4, Spectra MaxPlus 384

Spectrofluorimetr Sinco FS-2

Spectrophotometer Evolution 201

Dariusz Martynowski  PhD – Laboratory Manager

Katarzyna Niedzwiedzka – Process Engineer

Marta Małycha – Process Engineer

  • Vanderlinde E, Zhong S, Li G, Martynowski D, Grochulski P, Howard PS. (2014) ‘Functional assembly of the type two secretion system in Aeromona hydrophila involves direct interaction between the periplasmic domains of the assembly factor ExeB and the secretin ExeD’ PloS One, 9(7)
  • Martynowski D, Grochulski P, Howard PS. (2013) ‘Structure of a periplasmic domain of the EpsAB fusion protein of the Vibrio vulnificus type II secretion system.’ Acta Cryst. D69(2):142-9
  • Jin L, Martynowski D, Zheng S, Wada T, Xie W, Li Y. (2010) ‘Structural basis for hydroxycholesterols as natural ligands of orphan nuclear receptor RORgamma.’ Mol Endocrinol.24(5):923-9
  • Sorci L, Martynowski D, Rodionov DA, Eyobo Y, Zogaj X, Klose KE, Nikolaev EV, Magni G, Zhang H, Osterman AL. (2009) ’Nicotinamide mononucleotide synthetase is the key enzyme for an alternative route of NAD biosynthesis in Francisella tularensis.’ PNAS;106(9):3083-8
  • Li Y, Zhang J, Schopfer FJ, Martynowski D, Garcia-Barrio MT, Kovach A, Suino-Powell K, Baker PR, Freeman BA, Chen YE, Xu HE. (2008),’Molecular recognition of nitrated fatty acids by PPAR gamma.’ Nat Struct Mol Biol. 15(8):865-7
  • Li Y, Kovach A, Suino-Powell K, Martynowski D, Xu HE. (2008),’ Structural and biochemical basis for the binding selectivity of peroxisome proliferator-activated receptor gamma to PGC-1alpha.’  J Biol Chem. 283(27):19132-9
  • Rodionov DA, Li X, Rodionova IA, Yang C, Sorci L, Dervyn E, Martynowski D, Zhang H, Gelfand MS, Osterman AL. (2008),’Transcriptional regulation of NAD metabolism in bacteria: genomic reconstruction of NiaR (YrxA) regulon.’, Nucleic Acids Res. 36(6):2032-46.
  • Lee J, Michael AJ, Martynowski D, Goldsmith EJ, Phillips MA. (2007),’Phylogenetic diversity and the structural basis of substrate specificity in the beta/alpha-barrel fold basic amino acid decarboxylases.’, J Biol Chem. 282(37):27115-25
  • Bochkareva E*, Martynowski D*, Seitova A, Bochkarev A. (2006), ‘Structure of the origin-binding domain of simian virus 40 large T antigen bound to DNA’, EMBO J., 25(24), 5961-5969             *equal contribution
  • Martynowski D, Eyobo Y, Li T, Yang K, Liu A, Zhang H (2006), ‘Crystal Structure of a-Amino-b-Carboxymuconate-e-Semialdehyde Decarboxylase (ACMSD): Insight into the Active Site and Catalytic Mechanism of a Novel Decarboxylation Reaction’, Biochemistry 45(35), 10412-10421
  • Yang K, Eyobo Y, Brand LA, Martynowski D, Tomchick D, Strauss E, Zhang H (2006),’Crystal Structure of a Type III Pantothenate Kinase: Insight Into the Mechanism of an Essential Coenzyme A Biosynthetic Enzyme Universally Distributed in Bacteria’, J. Bacteriology 188(15), 5532-5540
  • Głowka ML, Martynowski D, Olczak A, Orlewska C, Foks H, Bojarska J, Szczesio M, Gołka J (2005), ‘Planarity of N′-(amino-2-pyridylmethylene)-hydrazide carbodithioic acid frame and crystal structure of its methyl ester dihydrate’, J. Chem. Cryst. 35 (6), 477-480
  • Grabowski SJ, Dubis AT, Martynowski D, Glowka ML, Palusiak M,Leszczynski J (2004), ’Crystal and molecular structure of pyrrole-2-carboxylic acid; electron delocalization of its dimmers – DFT and MP2 calculations’, J. Phys. Chem. A108(27), 5815-5822
  • Glowka ML, Martynowski D, Olczak A, Bojarska J, Szczesio M, Kozlowska K(2003), ‘Intramolecular hydrogen bond between 4-oxo and 3-carboxylic groups in quinolones and their analogs. Crystal structures of 7-methyl- and 6-fluoro-1,4-dihydro-4-oxocinnoline-3-carboxylic acids’, J. Mol. Struct., 658, 43-50
  • Stanczak A, Ochocki Z, Martynowski D, Glowka ML, Nawrot E (2003), ’Synthesis, structure and antibacterial activity of 4-imino-1,4-dihydrocinnoline-3-carboxylic acid derivatives as isosteric analogues of quinolones’, Arch. Pharm. Pharm. Med. Chem., 1, 18-30
  • Glowka ML, Olczak A, Martynowski D, Kozlowska K, Kulpinski J (2002), ‘Structural consequences of hindered rotation of tolyl substituent in 2,2,4,4,6,6-hexamethyl-1,3,5-triolylcyclotrisilazanes. Crystal structure of o-, m-, p-tolyl derivatives’, J. Mol. Struct., 613, 145-151
  • Orlewska C, Foks H, Sowinski P, Martynowski D, Olczak A, Glowka ML (2001), ‘Synthesis, structure and tuberculostatic activity of N’-(Amino-pyridyl-methylene)-hydrazinecarbodithioic Acid Methyl Esters’, Pol. J. Chem., 75, 1237
  • Glowka ML, Martynowski D, Napieraj A, Olczak A, Stanczak A, Ochocki Z, Lewgowd W (1999), ‘Structure of 1,4-dihydro-1-ethyl-4-iminecinnoline-3-carboxylic acids, classical isosters of quinolone antibacterials: Crystal structures of hydrochlorides of 7-chloro-and 7-methyl derivatives’, J. Chem. Cryst., 29, 687-693
  • Glowka ML; Martynowski D; Olczak A; et al. (1999), ‘Crystal and molecular structures of 1,1-bis(methylthio)-4-(2-pyridyl)-2,3,5-triaza-1,3-pentadiene and its 5-phenyl derivative’, Pol. J. Chem., 73, 845-851
  • Kaminski Z, Markowicz SW, Kolesinska B, Martynowski D, Glowka ML (1998), ‘Synthesis of Chiral 2,4-Dichloro-6-menthoxy-1,3,5-triazines and 2-Chloro-4,6-dimethoxy-1,3,5-triazines as Enantiodifferentiating Coupling Reagents. An X-Ray Study on 2,4,6-Trimethoxy-1,3,5-triazine’, Synth. Commun., 28, 2689-2696
  • Glowka ML, Martynowski D, Kozlowska K (1998), ‘Stacking of six membered aromatic rings in crystals’, Mol. Struct., 474, 81-89
  • Glowka ML, Olczak A, Martynowski D, Staszewska A (1997), ‘Quinoid Structure of Diazine Ring in 4-Iminecinnolines. Crystal and Molecular Structure of 3-Carbomoilo-1,4-dihydro-4-imino-1-methylcinnoline’, Pol. J. Chem., 71, 170-175
  • Krygowski TM, Howard ST, Martynowski D, Glowka ML (1997), ‘Imbalance of Kekule Structures in 2,4,6-trimethoxy-s-triazine’, J. Phys. Org. Chem., 10, 125-127
  • Kaminski Z, Glowka ML, Olczak A, Martynowski D (1996), ‘Thermal Isomerization of 2-Acyloxy-4,6-dimethoxy-1,3,5-triazines to 1-Acyl-3,5-dimethyl-1,3,5-triazin-2,4,6(1H,3H,5H)-triones. Crystal Structure of 1-(2,2-Dimethyloropionyloxy)-3,5-dimethyl-1,3,5-triazin-2,4,6-(1H,3H,5H)-triones’, Pol. J. Chem., 70, 1316-1323
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Posted by PORT - Polski Ośrodek Rozwoju Technologii, Posted on 10.02.2016
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