Biosynthesis of UDP-α-N-Acetyl-d-mannosaminuronic Acid and CMP-ß-N-Acetyl-d-neuraminic Acid for the Capsular Polysaccharides of Campylobacter jejuni.

Campylobacter jejuni is a human pathogen and a leading cause of food poisoning in North America and Europe. The exterior surface of the bacterial cell wall is attached to a polymeric coat of sugar molecules known as the capsular polysaccharide (CPS) that helps protect the organism from the host immune response. The CPS is composed of a repeating sequence of common and unusual sugar residues. In the HS:11 serotype of C. jejuni, we identified two enzymes in the gene cluster for CPS formation that are utilized for the biosynthesis of UDP-α-N-acetyl-d-mannosaminuronic acid (UDP-ManNAcA). In the first step, UDP-α-N-acetyl-d-glucosamine (UDP-GlcNAc) is epimerized at C2 to form UDP-α-N-acetyl-d-mannosamine (UDP-ManNAc). This product is then oxidized by a NAD+-dependent C6-dehydrogenase to form UDP-ManNAcA. In the HS:6 serotype (C. jejuni strain 81116), we identified three enzymes that are required for the biosynthesis of CMP-ß-N-acetyl-d-neuraminic acid (CMP-Neu5Ac). In the first step, UDP-GlcNAc is epimerized at C2 and subsequently hydrolyzed to form N-acetyl-d-mannosamine (ManNAc) with the release of UDP. This product is then condensed with PEP by N-acetyl-d-neuraminate synthase to form N-acetyl-d-neuraminic acid (Neu5Ac). In the final step, CMP-N-acetyl-d-neuraminic acid synthase utilizes CTP to convert this product into CMP-Neu5Ac. A bioinformatic analysis of these five enzymes from C. jejuni serotypes HS:11 and HS:6 identified other bacterial species that can produce UDP-ManNAcA or CMP-Neu5Ac for CPS formation.

Metabolic Engineering of Escherichia coli for High-Titer Biosynthesis of 3'-Sialyllactose.

3'-Sialyllactose (3'-SL) is among the foremost and simplest sialylated breast milk oligosaccharides. In this study, an engineered Escherichia coli for high-titer 3'-SL biosynthesis was developed by introducing a multilevel metabolic engineering strategy, including (1) the introduction of precursor CMP-Neu5Ac synthesis pathway and high-performance α2,3-sialyltransferase (α2,3-SiaT) genes into strain BZ to achieve de novo synthesis of 3'-SL; (2) optimizing the expression of glmS-glmM-glmU involved in the UDP-GlcNAc and CMP-Neu5Ac synthesis pathways, and constructing a glutamine cycle system, balancing the precursor pools; (3) analysis of critical intermediates and inactivation of competitive pathway genes to redirect carbon flux to 3'-SL biosynthesis; and (4) enhanced catalytic performance of rate-limiting enzyme α2,3-SiaT by RBS screening, protein tag cloning. The final strain BZAPKA14 yielded 9.04 g/L 3'-SL in a shake flask. In a 3 L bioreactor, fed-batch fermentation generated 44.2 g/L 3'-SL, with an overall yield and lactose conversion of 0.53 g/(L h) and 0.55 mol 3'-SL/mol, respectively.

Determinação do caseinomacropeptídeo em leite UAT por espectroscopia no infravermelho próximo e regressão por mínimos quadrados parciais / Determination of caseinomacropeptide in uht milk using near infrared spectroscopy and pls regression

O presente trabalho foi realizado com o objetivo de desenvolver um método para a determinação de CMP em leite UAT por meio da aplicação da espectroscopia de infravermelho próximo. Leites UAT de oito marcas diferentes foram utilizados para a construção dos modelos de calibração. Os resultados demonstram que, para o desenvolvimento de um modelo de calibração adequado para a determinação de CMP em leite UAT, deve-se utilizar, juntamente com o método de regressão PLS, o método de seleção de espectros máxima distância e os pré-tratamentos 2ª derivada e variável normal padronizada. Além disso, pôde-se determinar que as regiões do infravermelho próximo mais correlacionadas com os movimentos vibracionais dos aminoácidos presentes no CMP foram: 1100-1310; 1400-1430; 1490-1550; 1640-1680; 1780-1970; 2020-2100 e 2310-2350nm. Conclui-se que a espectroscopia de infravermelho próximo pode ser uma alternativa para a determinação de CMP em leite UAT, desde que haja um conjunto de calibração com amostras representativas da população a ser predita no futuro.(AU)

his work´s objective was to develop an UHT milk caseinomacropeptide determination method trough NIR spectroscopy application. Eight UHT milk trademarks are used to produce a mathematical calibration model. The results of NIR analysis suggested that to produce a suitable calibration model, partial least-square regression (PLSR) must be used, with maximum distance in wavelenght space to select spectra, pre - treatment with 2nd derivative and standard normal variant (SNV). Also, suitable near-infrared regions more correlated with CPM aminoacids vibrational movements: 1100-1310; 1400-1430; 1490-1550; 1640-1680; 1780-1970; 2020-2100; and 2310-2350nm. Therefore, NIR spectroscopy can be an alternative to caseinomacropeptide determination of UHT milk, since there was a representative calibration set with a large enough and representative sample of entire population to be predicted in the future.(AU)

Correlating mesophilic counts to the pseudo-CMP content of raw milk / Correlação entre contagem de mesófilos e índice de Pseudo-CMP em leite cru

A presente comunicação objetivou avaliar a quantificação do caseínomacropeptídeo (CMP), bem como diferenciá-lo (devido à adulteração com soro) do pseudo-CMP (devido à proteólise bacteriana) em amostras de leite cru coletadas nos domicílios do sul do Brasil. Os resultados reforçam a necessidade de práticas higiênicas durante a ordenha e estocagem do leite. As amostras de leite estudadas não estavam adulteradas por adição de soro, mostrando que a análise por cromatografia de exclusão por tamanho deve ser complementada a fim de revelar a identidade do peptídeo (CMP ou pseudo-CMP). A contagem bacteriana total (TBC) também se mostrou útil como indicador da contaminação do leite por micro-organismos proteolíticos, uma vez que uma relação diretamente proporcional entre TBC e pseudo-CMP foi estabelecida.(AU)

Estudio radomizado simple ciego mediante placebo efecto del citidin monofosfato-nucleo CMP(r) en la paralisi facial al frigore: Hospital de Clínicas, servicio de fisioterapia, año 2001 / CMP in the treatment of a frigore facil palsy: simpe randomized study

Pregunta de investigación. ¿Es más rápida la respuesta clínica favorable en el tratamiento de la Parálisi Facial a Frigore si incluimos el fármaco Núcleo CMP?. Hipótesis: En pacientes consultantes de Fisioterapía- Hospital de Clínicas gestión 2001, con Parálisis Facial a Frigore emplear como parte del tratamiento el fármaco Núcleo CMP, además del tratamiento porFisioterapia y corticoesteroide: resulta favorable en cuanto a su recuperación más rápida; frente a pacientes que no recibieron el medicamaento y solo se les dio placebo. Objetivos. 1 Determinar la rapidez en la mejoría clínica en pacientes con parálisi facial a frigore tratados en la unidad de Fisioterapía que recibieron el fármaco Núcleo CMP y aquellos que no lo recibieron. Demostrar el tiempo en el cual se presenta la respuesta favorable en abos grupos de pacientes con parálisis facial que recibieron el fármaco y los queno lo recibieron. Diseño. Ensayo Clínico, simple ciego, randomizado. Lugar. Hospital de Clínicas- La Paz- Unidad Medicina Familiar y Rehabilitación. Material y métodos. 1. Etapa. Obtención de la muestra y randomización pormedio dela tabla de números aleatorios delpaquete de estadística STATS TM, con demostración de homogeneidad. 2. Etapa Examén clínico neurológico, cálculo de medidas de efecto. Resultados. Se aleatorizaron a Núcleo CMP (n=27) y placebo (n=30) un total de 57 pacientes. Concluyeron el estudio 24 asignados a placebo y 20 tratados con núcleo CMP. Para la variable de eficacia principal (tiempo transcurrido hasta la mejoría neurológica confirmada), el análisis de las observaciones durante un seguimiento de 8 semanas/pacientes, indico de manifiesto una diferencia significativa entre ambos grupos; favorables al núcleo CMP (p=0.0026). Confirmado por la prueba X2 con aceptación de la Hipótesis del estudio. Esta evolución favorable se observó en (38.9 porciento) de los 2o pacientes asigandos al núcleo CMP (p= 0.039), que desarrollaron una progresión clínica favorable confirmada desde el 10mo día de tratamiento y en (34.8 por ciento) de 24 pacientes tratados con placebo. La evolución primaria se confirmó mediante examén neurológico. Conclusión. Se cumple la hipótesis del estudio, por ser el fármaco efectivo en la recuperación temprana de la Parálisis facial a frigore, "según el estudi", su uso sin embargo dependerá del criterio médico y la comparación por metaanálisis, con resultados de estudios más grandes.

Effect of the nucleotides CMP and UMP on exhaustion in exercise rats

The aetiology of muscle fatigue has yet not been clearly established. Administrationof two nucleotides, cytosine monophosphate (CMP) and uridine monophosphate(UMP), has been prescribed for the treatment of neuromuscular affections inhumans. Patients treated with CMP/UMP recover from altered neurological functionsand experience pain relief, thus the interest to investigate the possible effect ofthe drug on exhausting exercise. With such aim, we have determined, in exercised ratstreated with CMP/UMP, exercise endurance, levels of lactate, glucose and glycogen,and the activity of several metabolic enzymes such as, creatine kinase (CK), lactatedehydrogenase (LDH), and aspartate aminotransferase (AST). Our results show thatrats treated with CMP/UMP are able to endure longer periods of exercise (treadmillrun).Before exercise, muscle glucose level is significantly higher in treated rats, suggestingthat the administration of CMP/UMP favours the entry of glucose in themuscle. Liver glycogen levels remains unaltered during exercise, suggesting thatCMP/UMP may be implicated in maintaining the level of hepatic glycogen constantduring exercise. Lactate dehydrogenase and aspartate aminotransferase activity is significantlylower in the liver of treated rats. These results suggest that administrationof CMP/UMP enable rats to endure exercise by altering some metabolic parameters (AU)

No dipsonible

Efecto del núcleo CMP forte en 46 pacientes con paraparesia espástica progresiva: estudio randomizado y ciego / Effect of CMP forte nucleus in 46 patients with progressive spastic paraparesis: randomized and blind study

Idiopatic or HTLV-1 associated progressive spastic paraparesis does not have a clear etiology or treatment. To assess the effects of a medication containing cytidinmonophosphate, uridintriphosphate and vitamin B 12 in the treatment of progressive spastic. Patients with the disease were randomly assigned to receive the Nucleus CMP forte (containing dysodic cytidinmonophosphate 5 mg,trisodic uridintriphosphate 3 mg and hydroxicobalamin 2 mg) tid or placebo during 6 months. Gait, spasticity, degree of neurogenic bladder and somatosensitive evoked potentials were assessed during treatment. Forty six patients aged 25 to 79 years old were studied, 24 were female and 29 HTLV-1 positive. Twenty two were treated with the drug and the rest with placebo. Gait and spasticity improved in 7 of 22 patients receiving the drug and 1 of 24 receiving placebo (p<0.05). Neurogenic bladder improved in 10 of 22 receiving the drug and 4 patients treated with the drug and in two of seven treated with placebo. The medication caused a modest improvement in patients with progressive spastic paraparesis and was free of side effects