The in vivo rituximab effects in B cell malignancies are only partially understood. Here we analyzed in a large chronic lymphocytic leukemia (CLL) cohort (n = 80) the inter-patient variability in CLL cell count reduction within the first 24 h of rituximab administration in vivo, and a phenomenon of blood repopulation by malignant cells after anti-CD20 antibody therapy. Larger CLL cell elimination after rituximab infusion was associated with lower pre-therapy CLL cell counts, higher CD20 levels, and the non-exhausted capacity of complement-dependent cytotoxicity (CDC). The absolute amount of cell-surface CD20 molecules (CD20 density x CLL lymphocytosis) was a predictor for complement exhaustion during therapy. We also describe that a highly variable decrease in CLL cell counts at 5 h (88 %-2%) following rituximab infusion is accompanied in most patients by peripheral blood repopulation with CLL cells at 24 h, and in ∼20 % of patients, this resulted in CLL counts higher than before therapy. We provide evidence that CLL cells recrudescence is linked with i) CDC exhaustion, which leads to the formation of an insufficient amount of membrane attack complexes, likely resulting in temporary retention of surviving rituximab-opsonized cells by the mononuclear-phagocyte system (followed by their release back to blood), and ii) CLL cells regression from immune niches (CXCR4dimCD5bright intraclonal subpopulation). Patients with major peripheral blood CLL cell repopulation exhibited a longer time-to-progression after chemoimmunotherapy compared to patients with lower or no repopulation, suggesting chemotherapy vulnerability of CLL cells that repopulate the blood.
- MeSH
- chronická lymfatická leukemie krev farmakoterapie imunologie patologie MeSH
- cytotoxicita imunologická imunologie MeSH
- komplement imunologie MeSH
- lidé MeSH
- následné studie MeSH
- protinádorové látky imunologicky aktivní terapeutické užití MeSH
- rituximab terapeutické užití MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Burkholderia pseudomallei and Chromobacterium violaceum are bacteria of tropical and subtropical soil and water that occasionally cause fatal infections in humans and animals. Microbial lectins mediate the adhesion of organisms to host cells, which is the first phase in the development of infection. Here we report the discovery of two novel lectins from the above-mentioned bacteria - BP39L and CV39L. The crystal structures revealed that the lectins possess a seven-bladed β-propeller fold. Functional studies conducted on a series of oligo- and polysaccharides confirmed the preference of BP39L for mannosylated saccharides and CV39L for rather more complex polysaccharides with a monosaccharide preference for β-l-fucose. The presented data indicate that the proteins belong to a currently unknown family of lectins.
- MeSH
- bakteriální proteiny metabolismus MeSH
- Burkholderia pseudomallei metabolismus MeSH
- Chromobacterium metabolismus MeSH
- fukosa metabolismus MeSH
- lektiny metabolismus MeSH
- lidé MeSH
- monosacharidy metabolismus MeSH
- polysacharidy metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- antigeny CD20 metabolismus MeSH
- biologické markery MeSH
- buněčná cytotoxicita závislá na protilátkách imunologie MeSH
- chronická lymfatická leukemie farmakoterapie imunologie metabolismus patologie MeSH
- cytotoxicita imunologická MeSH
- fenotyp MeSH
- komplement imunologie MeSH
- lidé MeSH
- nádorové buňky kultivované MeSH
- nádorové mikroprostředí účinky léků imunologie MeSH
- protinádorové látky imunologicky aktivní farmakologie terapeutické užití MeSH
- receptory antigenů B-buněk metabolismus MeSH
- rituximab farmakologie terapeutické užití MeSH
- signální transdukce účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Histone modifications have a profound impact on the chromatin structure and gene expression and their correct establishment and recognition is essential for correct cell functioning. Malfunction of histone modifying proteins is associated with developmental defects and diseases and detailed characterization of these proteins is therefore very important. The lysine specific demethylase KDM2A is a CpG island binding protein that has been studied predominantly for its ability to regulate CpG island-associated gene promoters by demethylating their H3K36me2. However, very little attention has been paid to the alternative KDM2A isoform that lacks the N-terminal demethylation domain, KDM2A-SF. Here we characterized KDM2A-SF more in detail and we found that, unlike the canonical full length KDM2A-LF isoform, KDM2A-SF forms distinct nuclear heterochromatic bodies in an HP1a dependent manner. Our chromatin immunoprecipitation experiments further showed that KDM2A binds to transcriptionally silent pericentromeric regions that exhibit high levels of H3K36me2. H3K36me2 is the substrate of the KDM2A demethylation activity and the high levels of this histone modification in the KDM2A-bound pericentromeric regions imply that these regions are occupied by the demethylation deficient KDM2A-SF isoform.
- MeSH
- centromera metabolismus MeSH
- chromozomální proteiny, nehistonové metabolismus MeSH
- demetylace * MeSH
- doména Jumonji s histondemethylasami chemie metabolismus MeSH
- F-box proteiny chemie metabolismus MeSH
- heterochromatin metabolismus MeSH
- izoenzymy chemie metabolismus MeSH
- lidé MeSH
- MFC-7 buňky MeSH
- proteinové domény MeSH
- vazba proteinů MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Structurally different and functionally diverse natural compounds - antitumour agents pyrrolo[1,4]benzodiazepines, bacterial hormone hormaomycin, and lincosamide antibiotic lincomycin - share a common building unit, 4-alkyl-L-proline derivative (APD). APDs arise from L-tyrosine through a special biosynthetic pathway. Its generally accepted scheme, however, did not comply with current state of knowledge. Based on gene inactivation experiments and in vitro functional tests with recombinant enzymes, we designed a new APD biosynthetic scheme for the model of lincomycin biosynthesis. In the new scheme at least one characteristic in each of five final biosynthetic steps has been changed: the order of reactions, assignment of enzymes and/or reaction mechanisms. First, we demonstrate that LmbW methylates a different substrate than previously assumed. Second, we propose a unique reaction mechanism for the next step, in which a putative γ-glutamyltransferase LmbA indirectly cleaves off the oxalyl residue by transient attachment of glutamate to LmbW product. This unprecedented mechanism would represent the first example of the C-C bond cleavage catalyzed by a γ-glutamyltransferase, i.e., an enzyme that appears unsuitable for such activity. Finally, the inactivation experiments show that LmbX is an isomerase indicating that it transforms its substrate into a compound suitable for reduction by LmbY, thereby facilitating its subsequent complete conversion to APD 4-propyl-L-proline. Elucidation of the APD biosynthesis has long time resisted mainly due to the apparent absence of relevant C-C bond cleaving enzymatic activity. Our proposal aims to unblock this situation not only for lincomycin biosynthesis, but generally for all above mentioned groups of bioactive natural products with biotechnological potential.
- Publikační typ
- časopisecké články MeSH
In the biosynthesis of lincosamide antibiotics lincomycin and celesticetin, the amino acid and amino sugar units are linked by an amide bond. The respective condensing enzyme lincosamide synthetase (LS) is expected to be an unusual system combining nonribosomal peptide synthetase (NRPS) components with so far unknown amino sugar related activities. The biosynthetic gene cluster of celesticetin was sequenced and compared to the lincomycin one revealing putative LS coding ORFs shared in both clusters. Based on a bioassay and production profiles of S. lincolnensis strains with individually deleted putative LS coding genes, the proteins LmbC, D, E, F and V were assigned to LS function. Moreover, the newly recognized N-terminal domain of LmbN (LmbN-CP) was also assigned to LS as a NRPS carrier protein (CP). Surprisingly, the homologous CP coding sequence in celesticetin cluster is part of ccbZ gene adjacent to ccbN, the counterpart of lmbN, suggesting the gene rearrangement, evident also from still active internal translation start in lmbN, and indicating the direction of lincosamide biosynthesis evolution. The in vitro test with LmbN-CP, LmbC and the newly identified S. lincolnensis phosphopantetheinyl transferase Slp, confirmed the cooperation of the previously characterized NRPS A-domain LmbC with a holo-LmbN-CP in activation of a 4-propyl-L-proline precursor of lincomycin. This result completed the functional characterization of LS subunits resembling NRPS initiation module. Two of the four remaining putative LS subunits, LmbE/CcbE and LmbV/CcbV, exhibit low but significant homology to enzymes from the metabolism of mycothiol, the NRPS-independent system processing the amino sugar and amino acid units. The functions of particular LS subunits as well as cooperation of both NRPS-based and NRPS-independent LS blocks are discussed. The described condensing enzyme represents a unique hybrid system with overall composition quite dissimilar to any other known enzyme system.
The gene lmbB2 of the lincomycin biosynthetic gene cluster of Streptomyces lincolnensis ATCC 25466 was shown to code for an unusual tyrosine hydroxylating enzyme involved in the biosynthetic pathway of this clinically important antibiotic. LmbB2 was expressed in Escherichia coli, purified near to homogeneity and shown to convert tyrosine to 3,4-dihydroxyphenylalanine (DOPA). In contrast to the well-known tyrosine hydroxylases (EC 1.14.16.2) and tyrosinases (EC 1.14.18.1), LmbB2 was identified as a heme protein. Mass spectrometry and Soret band-excited Raman spectroscopy of LmbB2 showed that LmbB2 contains heme b as prosthetic group. The CO-reduced differential absorption spectra of LmbB2 showed that the coordination of Fe was different from that of cytochrome P450 enzymes. LmbB2 exhibits sequence similarity to Orf13 of the anthramycin biosynthetic gene cluster, which has recently been classified as a heme peroxidase. Tyrosine hydroxylating activity of LmbB2 yielding DOPA in the presence of (6R)-5,6,7,8-tetrahydro-L-biopterin (BH4) was also observed. Reaction mechanism of this unique heme peroxidases family is discussed. Also, tyrosine hydroxylation was confirmed as the first step of the amino acid branch of the lincomycin biosynthesis.
- MeSH
- antibakteriální látky biosyntéza MeSH
- bakteriální proteiny genetika metabolismus MeSH
- cirkulární dichroismus MeSH
- dihydroxyfenylalanin metabolismus MeSH
- Escherichia coli enzymologie genetika MeSH
- exprese genu MeSH
- hem chemie metabolismus MeSH
- hemoproteiny genetika metabolismus MeSH
- hydroxylace MeSH
- linkomycin biosyntéza MeSH
- multigenová rodina MeSH
- rekombinantní proteiny genetika metabolismus MeSH
- Streptomyces enzymologie genetika MeSH
- tyrosin-3-monooxygenasa genetika metabolismus MeSH
- tyrosin metabolismus MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- železo chemie metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Chemical diversity: Two SAM-dependent N-methyltransferases-LmbJ from the biosynthesis of the antibiotic lincomycin and CcbJ from celesticetin biosynthesis-have been characterized and compared. Both tested enzymes form multimers and are able to utilize N-demethyllincomycin, the natural substrate of LmbJ, with comparable efficiency.
- MeSH
- antibakteriální látky biosyntéza chemie MeSH
- biokatalýza * MeSH
- linkomycin biosyntéza chemie MeSH
- linkosamidy biosyntéza chemie MeSH
- methyltransferasy chemie metabolismus MeSH
- molekulární konformace MeSH
- substrátová specifita MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Článek popisuje úskalí péče o nemocnou s BMI nad 50 z pohledu ošetřovatelského personálu (sester). Jaké jsou možnosti péče v současných podmínkách oddělení JIP. Součástí příspěvku je také kazuistika morbidně obézní pacientky se zamyšlením nad spoluprací s rodinou a okolím nemocné.
- MeSH
- dietoterapie MeSH
- dospělí MeSH
- dýchací soustava MeSH
- hospitalizace MeSH
- hygiena MeSH
- hypotenze MeSH
- index tělesné hmotnosti MeSH
- lidé MeSH
- morbidní obezita * ekonomika komplikace terapie MeSH
- ošetřovatelská péče MeSH
- renální insuficience komplikace MeSH
- srdeční selhání MeSH
- výsledek terapie MeSH
- zdravotní sestry MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- kazuistiky MeSH
