RESUMEN
The stable strain of methylotrophic yeast Pichia pastoris secreting human serum albumin into cultural medium was obtained. Optimal conditions for expression of the protein were determined. We characterized the recombinant protein by mass spectrometry and circular dichroism and analyzed its catalytic activity.
Asunto(s)
Pichia , Proteínas Recombinantes/biosíntesis , Albúmina Sérica/biosíntesis , Catálisis , Dicroismo Circular , Humanos , Pichia/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Albúmina Sérica/química , Albúmina Sérica/genéticaRESUMEN
AIM: To study possible pathogenetic role and clinical significance of DNA-hydrolysing autoantibodies (autoAB) or DNA-abzymes in patients with rheumatoid arthritis. MATERIAL AND METHODS: Prevalence of DNA-abzymes and their catalytic activity were studied in 400 patients with rheumatoid arthritis (RA) and 88 healthy donors matched by age and gender. RESULTS: Associated with DNA-binding autoAB DNA-hydrolysing activity was detected in 41.5% cases of RA. DNA-abzymes were maximally active in men with rheumatoid factor (RF) and women without RF, while it was minimal in men without RF and women with RF. By catalytic activity there was no significant differences between patients with RF and without it. The highest catalytic activity of DNA abzymes was detected in patients with distinct extraarticular pathology. DNA-abzymes were also active in patients with x-ray stage III-IV of the disease in association with high prevalence of catalytic autoAB. DNA abzymes were also active in patients with RA activity stage II and III. CONCLUSION: It is possible to use DNA-abzymes in clinical practice for monitoring of the disease activity in RA.
Asunto(s)
Anticuerpos Catalíticos/sangre , Artritis Reumatoide/diagnóstico , ADN/inmunología , Monitorización Inmunológica , Anticuerpos Catalíticos/metabolismo , Artritis Reumatoide/diagnóstico por imagen , Artritis Reumatoide/inmunología , Autoanticuerpos/sangre , Autoanticuerpos/metabolismo , ADN/metabolismo , Progresión de la Enfermedad , Femenino , Humanos , Hidrólisis , Masculino , Pronóstico , Radiografía , Factor Reumatoide/sangreRESUMEN
AIM: To evaluate pathogenetic and clinical significance of autoantibodies (AAB) with catalytic activity in the serum of patients with autoimmune myocarditis (AM). MATERIAL AND METHODS: The study was made on the sera from 99 patients with AM of different course: malignant, benign, myocardiosclerosis (MCS). In addition to standard immunological parameters, the study was made of serum levels of anticardiomyosine-antiCM (protabzymes) and anti-DNA (DNA-abzymes) of AAB. After obtaining anti-CM and anti-DNA IgG-AT, we determined non-specific and specific proteolytic activity of anti-CM. RESULTS: Maximal specific activity of protabzymes was seen in 73% patients with malignant AM, it correlated with blood levels of anti-CM AAB, DNA-abzymes activity was very high in 45% patients. In MCS proteolytic activity of autoAT was absent in 61% patients. In benign AM occurrence of protabzymes was confirmed in 35% cases. Elevated DNA-hydrolyzing activity of DNA-abzymes occurred in 13% cases. The activity had no significant correlation with serum titers of AB. In MCS proteolytic activity of AAB was absent in 61% cases, but high activity of anti-CM AAB was in 28%. The activity of DNA-abzymes in 44% ranged considerably which, in seropositive cases, detected significant correlation with serum titers of DNA-binding autoAT. CONCLUSION: Evaluation of catalytic activity of AAB may be considered as a criterial test assessing the stage, clinical variants and severity of AM. It also permits formulation of the disease prognosis and its possible outcomes.
Asunto(s)
Anticuerpos Catalíticos/sangre , Autoanticuerpos/sangre , Enfermedades Autoinmunes/diagnóstico , Miocarditis/diagnóstico , Anticuerpos Catalíticos/metabolismo , Autoanticuerpos/genética , Enfermedades Autoinmunes/inmunología , Miosinas Cardíacas/inmunología , Catálisis , ADN/inmunología , Humanos , Inmunoglobulina G/genética , Miocarditis/inmunología , PronósticoRESUMEN
A method for expression of an onconase gene leading to a soluble form of the protein was developed. The enzymatic and cytotoxic properties of the protein's recombinant forms were studied. Recombinant onconase with an additional N-terminal Met residue isolated in nondenaturing conditions did not substantially differ from the native enzyme in ribonucleolytic activity. The addition of a 33-mer peptide containing auxiliary elements for the simplification of isolation and detection of the recombinant protein did not affect the enzyme properties of onconase. The method proposed is useful for the onconase structure-function relation studies and enables construction of onconase-based fusion proteins for anticancer therapy.