Evaluation of components for the heterologous expression of Mycobacterium chubuense NBB4 monooxygenases

Bacterial monooxygenases have important roles in the biogeochemical cycling of carbon and nitrogen, as well as potential applications in biocatalysis of pharmaceutically important epoxides, and bioremediation of chlorinated pollutants. However, use of monooxygenases for biotechnology requires understanding of microbial physiology and the catalytic properties (e.g. substrate range and kinetic parameters) of monooxygenases themselves. Heterologous expression of monooxygenases is one way to advance our fundamental knowledge of MO functions, and also to enable future applications. This project focused on designing a new heterologous expression system for monooxygenases, using the soluble di-iron ethene monooxygenase (EtnMO) and the particulate copper hydrocarbon monooxygenase (HMO) from Mycobacterium strain NBB4 as models. The new Mycobacterium/E. coli shuttle vector (pMycoFos) allows stable cloning of the EtnMO and HMO enzymes in E. coli, and functional expression of these in Mycobacterium smegmatis mc2-155. This study evaluated the requirement for accessory proteins such as chaperonins and catalase peroxidase in the hydrocarbon metabolism of strain NBB4, and charactersied three such proteins (Cpn60.1, Cpn10, and KatG) by expression and purification in E. coli. Although co-expression of these proteins did not enhance MO activity in M. smegmatis, evidence for a role for KatG in alkane metabolism was obtained from the higher peroxidase activity seen in butane-grown cells relative to cells grown on acetate. Codon-optimisation was tested as a means of enabling expression of the EtnMO genes in E. coli, but this approach was not successful. However, possible evidence for the importance of codon usage was obtained from experiments on heterologous expression of the EtnMO in Pseudomonas putida strain KT2440. This high-GC bacterium was capable of expression of the unmodified EtnMO genes. This is the first example of functional expression of an alkene monooxygenase (group 4 SDIMO) in a gram-negative organism

Genetics of hereditary head and neck paragangliomas

Item does not contain fulltextBACKGROUND: The purpose of this study was to give an overview on hereditary syndromes associated with head and neck paragangliomas (HNPGs). METHODS: Our methods were the review and discussion of the pertinent literature. RESULTS: About one third of all patients with HNPGs are carriers of germline mutations. Hereditary HNPGs have been described in association with mutations of 10 different genes. Mutations of the genes succinate dehydrogenase subunit D (SDHD), succinate dehydrogenase complex assembly factor 2 gene (SDHAF2), succinate dehydrogenase subunit C (SDHC), and succinate dehydrogenase subunit B (SDHB) are the cause of paraganglioma syndromes (PGLs) 1, 2, 3, and 4. Succinate dehydrogenase subunit A (SDHA), von Hippel-Lindau (VHL), and transmembrane protein 127 (TMEM127) gene mutations also harbor the risk for HNPG development. HNPGs in patients with rearranged during transfection (RET), neurofibromatosis type 1 (NF1), and MYC-associated factor X (MAX) gene mutations have been described very infrequently. CONCLUSION: All patients with HNPGs should be offered a molecular genetic screening. This screening may usually be restricted to mutations of the genes SDHD, SDHB, and SDHC. Certain clinical parameters can help to set up the order in which those genes should be tested. (c) 2013 Wiley Periodicals, Inc. Head Neck 36: 907-916, 2014

Update on primary head and neck mucosal melanoma

Primary mucosal melanomas (PMMs) of the head and neck are uncommon malignancies that arise mainly in the nasal cavity and paranasal sinuses, followed by the oral cavity. The mainstay of treatment is radical surgical resection followed by adjuvant radiotherapy in selected patients with high-risk features. Multimodality therapy has not been well studied and is not standardized. Adjuvant radiotherapy seems to improve locoregional control but does not improve overall survival (OS). Elective neck dissection is advocated in patients with oral PMM. Systemic therapy should be considered only for patients with metastatic or unresectable locoregional disease. Despite improvements in the field of surgery, radiotherapy, and systemic therapy, patients with PMM still face a very unfavorable prognosis (5-year disease-free survival [DFS] <20%) with high rates of locoregional recurrence and distant metastasis. The present review aims to summarize the current state of knowledge on the molecular biology, pathological diagnosis, and management of this disease. (c) 2015 Wiley Periodicals, Inc. Head Neck 38: 147-155, 2016

Reply to the letter to the editor by Straetmans et al

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Reply to the letter to the editor by Straetmans et al

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