RESUMEN
A hyperthermostable endoglucanase from Pyrococcus horikoshii with the capability of hydrolyzing crystalline cellulose was analyzed. A protein engineering study was carried out to obtain a reduced-size mutant. Five amino acid residues at both the N- and C-terminus were found to be removable without any loss of activity or thermal stability. Site-directed mutagenesis was also performed on R102, N200, E201, H297, Y299, E342, and W377, residues possibly involved in the active center or in the recognition and binding of a cellulose substrate. The activity of the resulting mutants was considerably decreased, confirming that the mutated residues were all important for activity. A reduced-size enzyme, as active as the wild-type endoglucanase, was successfully obtained, plus the residues critical for its activity and specificity were confirmed. Consequently, an engineered enzyme with a reduced size was obtained, and the amino acids essential for activity were confirmed by site-directed mutagenesis and comparison with a known three-dimensional structure.
Asunto(s)
Celulasas/genética , Celulasas/metabolismo , Pyrococcus horikoshii/enzimología , Sitios de Unión , Celulasas/química , Celulosa/metabolismo , Estabilidad de Enzimas , Mutagénesis Sitio-Dirigida , Pyrococcus horikoshii/genética , Eliminación de SecuenciaRESUMEN
PURPOSE: The National Health Insurance Service-Health Screening Cohort (NHIS-HEALS) is a cohort of participants who participated in health screening programmes provided by the NHIS in the Republic of Korea. The NHIS constructed the NHIS-HEALS cohort database in 2015. The purpose of this cohort is to offer relevant and useful data for health researchers, especially in the field of non-communicable diseases and health risk factors, and policy-maker. PARTICIPANTS: To construct the NHIS-HEALS database, a sample cohort was first selected from the 2002 and 2003 health screening participants, who were aged between 40 and 79 in 2002 and followed up through 2013. This cohort included 514 866 health screening participants who comprised a random selection of 10% of all health screening participants in 2002 and 2003. FINDINGS TO DATE: The age-standardised prevalence of anaemia, diabetes mellitus, hypertension, obesity, hypercholesterolaemia and abnormal urine protein were 9.8%, 8.2%, 35.6%, 2.7%, 14.2% and 2.0%, respectively. The age-standardised mortality rate for the first 2 years (through 2004) was 442.0 per 100 000 person-years, while the rate for 10 years (through 2012) was 865.9 per 100 000 person-years. The most common cause of death was malignant neoplasm in both sexes (364.1 per 100 000 person-years for men, 128.3 per 100 000 person-years for women). FUTURE PLANS: This database can be used to study the risk factors of non-communicable diseases and dental health problems, which are important health issues that have not yet been fully investigated. The cohort will be maintained and continuously updated by the NHIS.
Asunto(s)
Tamizaje Masivo/estadística & datos numéricos , Enfermedades no Transmisibles/epidemiología , Adulto , Anciano , Estudios Transversales , Bases de Datos Factuales , Femenino , Estudios de Seguimiento , Encuestas Epidemiológicas , Humanos , Masculino , Persona de Mediana Edad , Programas Nacionales de Salud/estadística & datos numéricos , Enfermedades no Transmisibles/mortalidad , Prevalencia , República de Corea/epidemiología , Factores de Riesgo , Enfermedades Estomatognáticas/epidemiologíaRESUMEN
A hyperthermophilic beta-1,4 endoglucanase (EGPh) from the hyperthermophilic archaeon Pyrococcus horikoshii exhibits a strong hydrolyzing activity toward crystalline cellulose. The characteristic features of EGPh are: (1) it appears to have disulfide bonds, which is rare among anaerobic hyperthermophilic archaeon proteins, and (2) it lacks a carbohydrate-binding domain, which is necessary for effective hydrolysis of cellulose. We first examined the relationship between the disulfide bonds and the catalytic activity by analyzing various cysteine mutations. The activities of the mutated enzymes toward carboxy methyl cellulose (CMC) increased without any loss in thermostability. Second, we prepared a fusion enzyme so that the thermostable chitin-binding domain of chitinase from P. furiosus was joined to the C-terminus of EGPh and its variants. These fusion enzymes showed stronger activities than did the wild-type EGPh toward both CMC and crystalline cellulose (Avicel).
Asunto(s)
Proteínas Arqueales/química , Celulasa/química , Quitinasas/química , Pyrococcus horikoshii/enzimología , Proteínas Recombinantes de Fusión/química , Proteínas Arqueales/genética , Catálisis , Celulasa/genética , Celulosa/química , Quitinasas/genética , Mutación Missense , Estructura Terciaria de Proteína/genética , Pyrococcus horikoshii/genética , Proteínas Recombinantes de Fusión/genéticaRESUMEN
Charcot-Marie-Tooth disease type 1A (CMT1A) is associated with duplication of chromosome 17p11.2-p12, whereas hereditary neuropathy with liability to pressure palsies (HNPP), which is an autosomal dominant neuropathy showing characteristics of recurrent pressure palsies, is associated with 17p11.2-p12 deletion. An altered gene dosage of PMP22 is believed to the main cause underlying the CMT1A and HNPP phenotypes. Although CMT1A and HNPP are associated with the same locus, there has been no report of these two mutations within a single family. We report a rare family harboring CMT1A duplication and HNPP deletion.