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The primary challenge to improving existing cancer treatment is to develop drugs that specifically target tumor cell. NGR peptide is tumor homing peptide that selectively target cancer cells while interleukin 15 is a pleiotropic cytokine with anticancer properties. This study computationally engineered a IL15-NGR fusion peptide by linking the homing peptide NGR with the targeting peptide IL-15. After evaluating and validating the chimeric peptide, we docked it to the IL-15Rα/IL-15Rß/γc heterodimer receptor, examining the interactions and binding energy and lastly, molecular dynamics simulations were performed. The secondary and tertiary structures, along with physicochemical properties of the designed IL-15-NGR chimeric protein, were predicted using GOR IV, trRosetta and ProtParam online servers respectively. The quality and 3D structure validation were confirmed via ProSA-web and SAVES 6.0 analysis which predicted an ERRAT score of 96.72%, with 97.6% of residues in the Ramachandran plot, validating its structure. Finally, Docking, MD simulations and interaction analysis were performed using ClusPro 2.0 and GROMACS and PDBsum, which exhibited significant hydrogen bonding and salt bridges, confirming the formation of a stable docked complex. These results were further corroborated by simulation analysis, which demonstrated a stable and dynamic behavior of the docked complex in a biological environment. The predicted high expression value of fusion protein was 0.844 in E.coli using SOLUPROT tool. These findings suggest efficient expression of the IL15-NGR fusion protein if its gene is inserted into E. coli and indicates its potential as a safe and effective anticancer treatment, paving the way for targeted therapeutic interventions.
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The aim of the current study was to explore the potential of human plasma-derived exosomes as versatile carriers for drug delivery by employing various active and passive loading methods. Exosomes were isolated from human plasma using differential centrifugation and ultrafiltration method. Drug loading was achieved by employing sonication and freeze thaw methods, facilitating effective drug encapsulation within exosomes for delivery. Each approach was examined for its effectiveness, loading efficiency and ability to preserve membrane stability. Methotrexate (MTX), a weak acid model drug was loaded at a concentration of 2.2 µM to exosomes underwent characterization using various techniques such as particle size analysis, transmission electron microscopy and drug loading capacity. Human plasma derived exosomes showed a mean size of 162.15 ± 28.21 nm and zeta potential of -30.6 ± 0.71 mV. These exosomes were successfully loaded with MTX demonstrated a better drug encapsulation of 64.538 ± 1.54 % by freeze thaw method in comparison 55.515 ± 1.907 % by sonication. In-vitro drug release displayed 60 % loaded drug released within 72 h by freeze thaw method that was significantly different from that by sonication method i.e., 99 % within 72 h (p value 0.0045). Moreover, cell viability of exosomes loaded by freeze thaw method was significantly higher than that by sonication method (p value 0.0091) suggested that there was membrane disruption by sonication method. In conclusion, this study offers valuable insights into the potential of human plasma-derived exosomes loaded by freeze thaw method suggest as a promising carrier for improved drug loading and maintenance of exosomal membrane integrity.
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Background and Aims: It is essential to have proper treatment and management for asthma in order to minimise symptoms, lessen the burden, and lower the chance of exacerbations. To better control asthma, the purpose of this study was to evaluate and enhance paediatric registrars' understanding and application of asthma treatment. Methods: The Sudan Medical Specialisation Board (SMSB) paediatric registrars provided data for this quasi-interventional study between April and September of 2021. Twice, both before to and following the intervention education sessions, the questionnaire was delivered. SPSS version 28 was used to analyse the data after it had been cleaned up in an Excel document. Results: 203 (or 77.8%) of the 261 were women. A substantial rise from 8.1 ± 4.12 SD to 18 ± 5.03 SD in the mean overall knowledge score of registrars between the pre-and post-intervention periods. A significant difference (p=0.001) was observed in the post-intervention phase, with first-year registrars (R1) demonstrating higher scores than their senior colleagues. The registrars' total knowledge scores did not differ significantly from one another during the pre-intervention period. The Global Initiative of Asthma (GINA) guidelines of management were implemented to a certain extent, according to the study. Of the registrars, 148 (56.7%) and 203 (77.8%) evaluated step one management in children ages 5 and under; 66 (25.3%) and 213 (81.6%) evaluated step one management in children ages 6 to 11; and 66 (25.3%) and 213 (81.6%) evaluated step one management in children ages 6 to 11 in pre- and post-intervention, respectively. Conclusion: Given that the intervention in this study greatly increased registrars' knowledge, doctors should obtain training on the GINA 2019 recommendations through conferences, workshops, and academic programmes. To find out why R1 outperforms their older counterparts, more investigation has to be done.
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Cancer continues to be leading cause of morbidity and mortality despite decades of research and advancement in chemotherapy. Most tumors can be reduced via standard oncology treatments, such as chemotherapy, radiotherapy, and surgical resection, and they frequently recur. Significant progress has been made since targeted cancer therapy inception in creation of medications that exhibit improved tumor-selective action. Particularly in preclinical and clinical investigations, fusion proteins have shown strong activity and improved treatment outcomes for a number of human cancers. Synergistically combining many proteins into one complex allows the creation of synthetic fusion proteins with enhanced characteristics or new capabilities. Signal transduction pathways are important for onset, development, and spread of cancer. As result, signaling molecules are desirable targets for cancer therapies, and significant effort has been made into developing fusion proteins that would act as inhibitors of these pathways. A wide range of biotechnological and medicinal applications are made possible by fusion of protein domains that improves bioactivities or creates new functional combinations. Such proteins may function as immune effectors cell recruiters to tumors or as decoy receptors for various ligands. In this review article, we have outlined the standard methods for creating fusion proteins and covered the applications of fusion proteins in treatment of cancer. This article also highlights the role of fusion proteins in targeting the signaling pathways involved in cancer for effective treatment.
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Antineoplásicos , Neoplasias , Humanos , Neoplasias/tratamiento farmacológico , Biotecnología , Antineoplásicos/farmacología , Antineoplásicos/uso terapéuticoRESUMEN
CONTEXT: Epidermal growth factor receptor (EGFR), a member of the HER receptor family, is over expressed in various cancer cells. Using tumor-specific antibodies to deliver cytotoxic agents directly to the tumor cells is an effective treatment strategy. Targeted therapy by fusing anti-EGFR scFv with tumor-specific cytokines promises the emergence of a new era. METHODS: We designed a novel immuno-apoptotic fusion protein, anti-EGFR scFv-IL-24, consisting of a specific cancer cell targeting antibody and recombinant cytokine IL-24 to explore its anti-cancerous potential. Amino acid sequences of both anti-EGFR scFv and IL-24 were fused using a specific rigid linker. In silico characterization of the designed fusion protein like to predict the primary, secondary, physiochemical properties, quality, and structural validation using online bioinformatic tools. The newly designed fusion protein consists of 402 amino acids that showed good quality with a predicted value of 76.7% having 81.5% residues in the most favored region as predicted by ERRAT2 and Ramachandran plot analysis. Docking and simulation studies were performed using HDOCK and Desmond module of Schrodinger. All the parameters of quality, validity, interaction analysis, and stability suggested that the fused molecule is fully operational and functional. The results of the study support that the anti-EGFR scFv-IL-24 fused protein could be proved as a novel candidate to combat cancer.
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Aminoácidos , Apoptosis , Secuencia de Aminoácidos , Biología Computacional , Simulación por ComputadorRESUMEN
Exosomes have recently been considered ideal biotherapeutic nanocarriers that broaden the frontiers of current drug delivery systems to overcome the shortcomings associated with cytokine-based immunotherapy. Using this approach, the current study aimed to assess anti-proliferative activity of purified IL-29 and exosomes encapsulated IL-29. The IL-29+pET-28a construct was transformed into Rosetta 2(DE3) cells which was used for the large-scale production of IL-29. Exosomes isolated from H1HeLa, and SF-767 cells using Total Exosome Isolation reagent were loaded with IL-29 via sonication. Isolation of exosomes was validated using their core protein signature by western blotting and specific miRNA profiles by RT-PCR. The drug loading efficiency of exosomes derived from H1HeLa cells was higher than that of SF-767-derived exosomes. The drug release kinetics of IL-29 encapsulated exosomes exhibited stable release of the recombinant drug. Around 50% of all cancer cell lines survived when IL-29 was administered at a concentration of 20 µg/mL. A survival rate of less than 10% was observed when cells were treated with 20 µg/mL IL-29 loaded exosomes. It was concluded that IL-29 loaded exosomes had a more significant cytotoxic effect against cancer cells, which might be attributed to sustained drug release, improved half-life, superior targeting efficacy, capacity to harness endogenous intracellular trafficking pathways, and heightened biocompatibility of exosomes.
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Antineoplásicos , Exosomas , Exosomas/metabolismo , Sistemas de Liberación de Medicamentos , Antineoplásicos/farmacología , Antineoplásicos/metabolismo , Citocinas/metabolismo , Factores Inmunológicos , Interleucinas/genética , Interleucinas/farmacología , Interleucinas/metabolismoRESUMEN
Azurin is a natural protein produced by Pseudomonas aeruginosa that exhibits potential anti-tumor, anti-HIV, and anti-parasitic properties. The current study aimed to investigate the potential of azurin protein against breast cancer using both in silico and in vitro analyses. The amino acid sequence of Azurin was used to predict its secondary and tertiary structures, along with its physicochemical properties, using online software. The resulting structure was validated and confirmed using Ramachandran plots and ERRAT2. The mature azurin protein comprises 128 amino acids, and the top-ranked structure obtained from I-TASSER was shown to have a molecular weight of 14 kDa and a quality factor of 100% by ERRAT2, with 87.4% of residues in the favored region of the Ramachandran plot. Docking and simulation studies of azurin protein were conducted using HDOCK and Desmond servers, respectively. The resulting analysis revealed that Azurin docked against p53 and EphB2 receptors demonstrated maximum binding affinity, indicating its potential to cause apoptosis. The recombinant azurin gene was successfully cloned and expressed in a BL21 (DE3) strain using a pET20b expression vector under the control of the pelB ladder, followed by IPTG induction. The azurin protein was purified to high levels using affinity chromatography, yielding 70 mg/L. In vitro cytotoxicity assay was performed using MCF-7 cells, revealing the significant cytotoxicity of the azurin protein to be 105 µg/mL. These findings highlight the potential of azurin protein as an anticancer drug candidate.
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Targeted delivery of therapeutic anticancer chimeric molecules enhances the efficacy of drug by improving cellular uptake and circulation time. Engineering the molecules to facilitate the specific interaction between chimeric protein and its receptor is critical to elucidate biological mechanism as well as accuracy in modeling of complexes. A theoretically designed novel protein-protein interfaces can serve as a bottom-up method for comprehensive understanding of interacting protein residues. This study was aimed for in silico analyses of a chimeric fusion protein against breast cancer. The amino acid sequences of the interleukin 24 (IL-24) and LK-6 peptide were used to design the chimeric fusion protein via a rigid linker. The secondary and tertiary structures along with physicochemical properties by ProtParam and solubility were predicted using online software. The validation and quality of the fusion protein was confirmed by Rampage and ERRAT2. The newly designed fusion construct has a total length of 179 amino acids. The top-ranked structure from alpha fold2 showed 18.1 KD molecular weight by ProtParam, quality factor of 94.152 by ERRAT, and a valid structure by a Ramachandran plot with 88.5% residues in the favored region. Finally, the docking and simulation studies were performed using HADDOCK and Desmond module of Schrodinger. The quality, validity, interaction analysis, and stability of the fusion protein depict a functional molecule. The fusion gene IL24-LK6 after cloning and expression in a suitable prokaryotic cell might be a useful candidate for developing a novel anticancer therapy.
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OBJECTIVES: Osteoarthritis (OA) causes long-term pain and disorders of lower extremities. Paracetamol is the drug of choice; however, NSAIDs, opioids and steroids are frequently employed in the symptomatic relief of OA. The prescribing of multiple analgesics leads to potential drug-drug interactions (pDDIs). The primary objective of this study was to identify the prevalence and predictors of pDDIs in OA. MATERIALS AND METHODS: A total of three hundred and eighty-six patients, either newly diagnosed or with a history of OA, were enrolled for this cross-sectional study. Data regarding patients' demographics, clinical characteristics and prescribed medications were recorded from the prescriptions and examined for the pDDIs using Medscape multidrug interaction checker. RESULTS: Among 386, most patients were females (53.4%). The most prevalent diagnoses were knee OA (39.7%) and unspecified OA (31.3%). Paracetamol and topical NSAIDs were underprescribed whereas an oral NSAID, Diclofenac, was the most frequently used drug in OA. Total of 109 pDDIs were found in 386 prescriptions, of which 63.3% DDIs were categorised as moderate, followed by minor (34.9%) and major (1.8%). CONCLUSIONS: This study finds a prevalence of DDIs and polypharmacy among the OA patients. Collaborative efforts among healthcare providers, pharmacists, and patients themselves are essential to optimize medication regimens and minimize the polypharmacy including the associated risks as well as DDIs.
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Ortopedia , Osteoartritis de la Rodilla , Femenino , Humanos , Masculino , Acetaminofén/efectos adversos , Estudios Transversales , Antiinflamatorios no Esteroideos/efectos adversos , Osteoartritis de la Rodilla/tratamiento farmacológico , Osteoartritis de la Rodilla/epidemiología , Osteoartritis de la Rodilla/inducido químicamente , Interacciones FarmacológicasRESUMEN
Cancer is a complicated, adaptable, and heterogeneous disease caused by a wide variety of genetic changes that might impair ability of cells to function normally. The majority of the tumors can only be shrunk using conventional oncology therapies like chemotherapy, radiation, and surgical resection, and the tumor often recurs. The inability of conventional cancer therapies to completely destroy the Cancer Stem Cells (CSCs) that otherwise lead to therapy resistance is thus addressed by therapeutic approaches that concentrate on targeting CSCs and their micro-environmental niche. In this review, we summarize approaches that are used for the development of fusion proteins and their therapeutic applications for treating cancer. The main purpose of making advancements towards the fusion technology instead of using conventional treatment methods is to achieve a prolonged half-life of the therapeutic drugs. The fusion of drugs to the immune response enhancing cytokines or the fusion of antibody and cytokines not only increases half-life but also increase the stability of the anti-tumor drug. Several molecules including different fragments of antibodies, cytokines, Human Serum Albumin, transferrin, XTEN polymers, Elastin-like polypeptides (ELPs) can be employed as a fusion partner and the resulting fusion proteins are reported to show enhanced anti-tumor response.
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Antineoplásicos , Neoplasias , Humanos , Péptidos/química , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Antineoplásicos/química , Polímeros/química , Neoplasias/tratamiento farmacológico , Tecnología , CitocinasRESUMEN
The objective of the study was to develop PEGylated protamine letrozole nanoparticles to combat human breast cancer by modifying the release pattern of letrozole. Breast cancer is amongst the most prevalent diseases in women due to overactivity of human epidermal growth factor receptor 2 (HER2). PEG-protamine letrozole nanoparticle formulation was designed and optimized to alter the release pattern of the drug. The size, morphology, and structure of PEG-protamine letrozole NP were characterized by FTIR, XRD, Zetasizer, and SEM analysis. The result showed the PEG-protamine letrozole nanoparticles were irregular in shape and have size ranging from 258 nm to 388 nm, polydispersity index 0.114 to 0.45, zeta potential of 11.2 mV, and entrapment efficiency 89.93%. XRD studies have confirmed that the crystal structure of letrozole has become amorphous. The drug release study maintained the prolonged release for 72 hours. Moreover, the PEG-protamine letrozole NPs displayed a strong anticancer action compared to MCF-7 cells with an IC50 70 µM for letrozole and 50 µM for PEG-protamine letrozole NPs. Overall, our results indicate that letrozole PEG-protamine NPs alter the release profile of letrozole, which could be an excellent approach for overcoming letrozole resistance in human breast cancer.
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Neoplasias de la Mama , Nanopartículas , Neoplasias de la Mama/tratamiento farmacológico , Portadores de Fármacos/química , Femenino , Humanos , Letrozol/farmacología , Letrozol/uso terapéutico , Células MCF-7 , Nanopartículas/química , Tamaño de la Partícula , Polietilenglicoles/química , Protaminas/química , Protaminas/farmacología , Protaminas/uso terapéuticoRESUMEN
Various types of cancer pose a notable threat to human health globally. To date, many researchers have undertaken the search for anticancer therapies. However, many anticancer therapeutic approaches accompany many undesirable hazards. In this respect, extracellular vesicles as a whole gained excessive attention from the research community owing to their remarkable potential for delivery of anticancer agents since they are involved in distal intercellular communication via biological cargoes. With the discovery of the fact that tumor cells discharge huge quantities of EVs, new insights have been developed in cancer diagnosis and treatment. Tumor-derived extracellular vesicles (TD-EVs) can be distinguished from the normal cell-derived EVs due to the presence of specific labels on their surface. TD-EVs carry specific oncogenic proteins and the nucleic acids on their surface membrane that participate in tumor progression. Moreover, the proportion of these nucleic acids and the protein greatly varies among malignant and healthy cell-derived EVs. The diagnostic potential of TD-EVs can be implied for the more precise and early-stage detection of cancer that was impossible in the past. This review examines the recent progress in prognostic, diagnostic, and therapeutic potential of the EVs derived from the tumor cells.
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Lectins are the oligomeric sugar-specific glycoprotein of nonimmune origin, are involved in the multiple biological recognition process, and have the capacity to perform a wide variety of physiological functions including antifungal, antiviral, antitumor, and cell agglutination. The main objective of the current study was to prepare lectin protein-loaded chitosan-TPP nanoparticles via ionic gelation methods with different CS/TPP ratios and to investigate anticancer potential against HepG2 cells. The best ratio showed the mean particle size (298.10 ± 1.9 nm, 21.05 ± 0.95 mv) with optimal encapsulation efficiencies of 52.435 ± 0.09%. The cytotoxicity was evaluated against HepG2 cells, and IC50 values obtained were 265 µg/ml for lectin protein and 105 µg/ml for lectin-loaded chitosan-TPP nanoparticles, respectively. The mRNA expression of proliferation markers like GPC3 was significantly decreased in hepatocellular carcinoma cells (HepG2) during lectin protein-loaded chitosan-TPP nanoparticle treatment. Apoptotic genes that indicating a marked increase in expression are Caspase 3, p53, and Bax, while Bcl2 and AFP showed a downregulation of expression after treatment of HepG2 cells with lectin-loaded chitosan-TPP nanoparticles. The preliminary findings of our study highlighted that lectin protein-loaded chitosan-TPP nanoparticles could be a promising anticancer agent.
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Antineoplásicos Fitogénicos/farmacología , Quitosano/análogos & derivados , Composición de Medicamentos/métodos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Lectinas/farmacología , Lepidium sativum/química , Nanopartículas/química , Antineoplásicos Fitogénicos/química , Antineoplásicos Fitogénicos/aislamiento & purificación , Apoptosis/efectos de los fármacos , Apoptosis/genética , Caspasa 3/genética , Caspasa 3/metabolismo , Quitosano/química , Portadores de Fármacos , Geles , Células Hep G2 , Humanos , Concentración 50 Inhibidora , Lectinas/química , Lectinas/aislamiento & purificación , Nanopartículas/ultraestructura , Tamaño de la Partícula , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Transducción de Señal , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , alfa-Fetoproteínas/genética , alfa-Fetoproteínas/metabolismo , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismoRESUMEN
In the original publication, one of the author names was missed in the author group.
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INTRODUCTION: Pakistan has one of the highest burdens of pneumococcal diseases in the world, but unfortunately studies in this demanding research area are limited in the region. Pneumococcal surface protein A (PspA) is the next generation pneumococcal vaccine candidate as the protein locates on the Streptococcus pneumoniae surface. Its gene, pspA, might be encoded by all pneumococci, and the protein has proven immunogenicity. The molecular characterization of PspA, pneumococcal serotype distribution and antibiotic susceptibility are important for regional diversity studies. METHODS: In this study, we examined 38 pneumococcal isolates from pneumococcal diseased (pneumonia/meningitis) patients blood or cerebrospinal fluid. There were no specific inclusion or exclusion criteria, but all the individuals [ages 1 month to 12 years (male/female)] had undergone no antibiotic treatment in at least the past 3 months and had no vaccination history. We investigated the serotype distribution, antibiotic susceptibility, prevalence of the PspA family and its active domain's fusion, expression and antigenicity. RESULTS: Our finding shows that serotype 19F is the most prevalent (23.6%) followed by 18B (15.78%) (non-vaccine type) in all isolated pneumococcal strains. All strains were susceptible to chloramphenicol and linezolid, while 80% were resistant to gentamycin. Genotyping revealed that ~ 80% (N = 31/38) of pneumococcal strains produce PspA belonging to family 2 and clade 3. We further selected three active domains of PspA (family 2 and clade 3) by in silico analysis, merged together into a fusion gene for expression study, and its antigenicity was analyzed by Western blotting. CONCLUSION: Serotypes 19F and 18B (non-vaccine type) are the most prevalent in the Pakistani pneumococcal isolates. The PspA family 2 proteins produced by Pakistani pneumococcal isolates have high sequence homologies with each other and differ from those produced by strains isolated in the rest of the world. The PspA fusion peptide had a proven antigenic response in western blotting, with no considerable correlation among pneumococcal serotypes, antibiotic susceptibility and PspA family/clade distribution.
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Interleukin-6 a pleiotropic cytokine involved in a wide range of biological activities. So the large-scale production of biologically active recombinant human interleukin-6 is important for its structural and functional studies. Here, we report an optimized method for shake flask fermentation and a simplified high-yield purification procedure for the recombinant interleukin-6. This high-yield expression method not only involves the optimization of the fermentation condition but also the single step purification method as well as a two-step denaturing and one-step refolding process. This approach replaces the more conventional procedure of protein solubilization and refolding. Through applying these strategies, the final cell density and overall product yield of the recombinant human interleukin-6 were obtained as 20.4 g as cell biomass and 150 mg as purified active protein from the I-L of the culture. The purified protein was characterized by HPLC and SDS-PAGE. The results of the current work demonstrate that the described method may be used to develop the process for industrial-scale production of the biologically active recombinant interleukin-6 protein.
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Fermentación , Interleucina-6/biosíntesis , Línea Celular Tumoral , Proliferación Celular , Relación Dosis-Respuesta a Droga , Humanos , Interleucina-6/aislamiento & purificación , Interleucina-6/metabolismo , Replegamiento Proteico , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismoRESUMEN
ABSTRACT Rapid advances in medicine and biotechnology resulted in the development of non-invasive diagnostic and prognostic biomarkers enabling convenient and accurate detection. Exosomes has recently emerged as non-invasive biomarker for a number of diseases including cancer. Exosomes are the small endosome originated membranous vesicles secreted in a number of biological fluids such as serum, saliva, urine, ascites, cerebrospinal fluid, etc. Exosomes contain microRNA proteins and mRNA which can be used as disease specific biomarkers. Here we reviewed recent advancement in the field of exosomes as diagnostic biomarker for cancer along with a brief overview of their biogenesis, function and isolation.
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Recent advancement in fermentation technologies resulted in the increased yields of recombinant proteins of biopharmaceutical and medicinal importance. Consequently, there is an important task to develop simple and easily scalable methods that can facilitate the production of high-quality recombinant protein. Most of the recent reports described the expression of recombinant human IL-1 receptor antagonist (rhIL-1Ra) in Escherichia coli using isopropyl-ß-d-thiogalacto pyranoside (IPTG), a nonmetabolizable and expensive compound, as an expression inducer. In this study, we describe the expression and one-step purification of gallbladder-derived rhIL-1Ra by autoinduction in E. coli. This method includes special media that automatically induce the target protein expression from T7 promoter and allow the production of the target protein in high yield than the conventional IPTG induction method. In addition to fermentation process improvements, one-step purification strategy is essential to make the process economical. We developed a single-step cation exchange chromatography and obtained 300 mg/L of rhIL-1Ra with 98% purity. Purified protein was characterized by SDS-PAGE and Ion exchange HPLC (IEX-HPLC). The described method can be used to scale up the production of rhIL-1Ra and other recombinant proteins.
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Expresión Génica , Proteína Antagonista del Receptor de Interleucina 1/biosíntesis , Proteína Antagonista del Receptor de Interleucina 1/química , Proteína Antagonista del Receptor de Interleucina 1/aislamiento & purificación , Escherichia coli/química , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Proteína Antagonista del Receptor de Interleucina 1/genética , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificaciónRESUMEN
The development of biomarkers of reproductive medicine is still in its infancy because many black boxes are still present in reproductive medicine. Novel approaches to human infertility diagnostics and treatment must be developed because reproductive medicine has lagged behind in the implementation of biomarkers in clinical medicine. Despite the dearth of the available literature, the current rapid pace of publications suggests that this gap will soon be filled therefore; this review is a précis of the research that has been done so far and will provide a basis for the development of biomarkers in reproductive medicine.
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Recombinant consensus interferon (CIFN) is a therapeutic protein with molecular weight of 19.5 kDa having broad spectrum antiviral activity. Recombinant human CIFN (rhCIFN) has previously been expressed in Escherichia coli using isopropyl-ß-d-thiogalactopyranoside (IPTG), a non-metabolizable and expensive compound, as inducer. For economical and commercial-scale recombinant protein production, it is greatly needed to increase the product yield in a limited time frame to reduce the processing cost. To reduce the cost of production of rhCIFN in E. coli, induction was accomplished by using lactose instead of IPTG. Lactose induction (14 g/L) in shake flask experiment resulted in higher yield as compared with 1 mM IPTG. Finally, with single-step purification on DEAE sepharose, 150 mg/L of >98% pure rhCIFN was achieved. In the present study, an attempt was made to develop a low cost process for producing quality product with high purity. Methods devised may be helpful for pilot-scale production of recombinant proteins at low cost.