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National Repository of Grey Literature 2 records found  Search took 0.01 seconds. 
Recombinant preparation of DNA binding domain of transcription factor TEAD4
 Zákopčaník, Marek ; Novák, Petr (advisor) ; Šulc, Miroslav (referee)
6 Abstract Transcription factors play a key role in the management of cell growth and differ- entiation and their deregulation is associated with many cancers. TEAD proteins utilise highly conserved DNA binding domain to recognise specific DNA sequences. This domain could facilitate new drug design and development. The goal of this master thesis includes recombinant preparation of DNA binding domain of transcriptional factor TEAD4 extended by a part of an unstruc- tured variable sequence, which connects this domain with transactivation domain. Purification steps include affinity chromatography followed by size exclusion chro- matography. The characterization of produced protein was performed by mass spectrometry and finally, native gel electrophoresis was used to prove the ability of the produced protein to bind DNA. During purification steps, a fragmentation from C-terminus was observed. Based on analysis of the mass spectra, three most represented forms of produced protein were described all of which were fragmented. The most abundant form (55%) consisted of amino acids 30-131 from TEAD4 protein. Second most abun- dant form (18%) consisted of amino acids 30-144 and the third form consisted of amino acids 30-81. Native gel electrophoresis verified the ability to bind DNA, the efficiency was however lower...
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Study of lectin-antibody interaction by using surface plasmon resonance
 Zákopčaník, Marek ; Hodek, Petr (advisor) ; Nosková, Libuše (referee)
Cystic fibrosis is an autosomal recessive disease affecting mostly Caucasian race. The disease is caused by mutations in CFTR gene. The most affected organ system is respiratory tract, because the changes lead to insufficient natural defence function of lungs, causing the lungs being susceptible to bacterial infections, which are the most common cause of death of CF patients. Among these bacteria is also Burkholderia cepacia, which causes rapid degradation of pulmonary tissue. Eradication of this bacterial infection is complicated by a number of antibiotic resistances and biofilm formation. Chicken antibodies against bacterial lectins are potential means of preventing bacterial lung infection of patients with CF. Using the affinity chromatography with BC2L-A immobilized column a specific antibody fraction was obtained. The immunoreactivity of the mixed antibodies, affinity purified antibodies and unbounded fraction was compared by ELISA. The comparison of immunoreactivity has emerged that the concentration of the specific antibodies has grown more than 10 times. Interaction of lectin-antibody was studied with SPR method. In the first arrangement, the binding strength of the active lectin to glycosylation of the unbounded fraction antibodies was measured. The dissociation constant of this...
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