Engineering a Lysine-ON Riboswitch for Metabolic Control of Lysine Production in Corynebacterium glutamicum.

Riboswitches are natural RNA elements that regulate gene expression by binding a ligand. Here, we demonstrate the possibility of altering a natural lysine-OFF riboswitch from Eschericia coli (ECRS) to a synthetic lysine-ON riboswitch and using it for metabolic control. To this end, a lysine-ON riboswitch library was constructed using tetA-based dual genetic selection. After screening the library, the functionality of the selected lysine-ON riboswitches was examined using a report gene, lacZ. Selected lysine-ON riboswitches were introduced into the lysE gene (encoding a lysine transport protein) of Corynebacterium glutamicum and used to achieve dynamic control of lysine transport in a recombinant lysine-producing strain, C. glutamicum LPECRS, which bears a deregulated aspartokinase and a lysine-OFF riboswitch for dynamic control of the enzyme citrate synthase. Batch fermentation results of the strains showed that the C. glutamicum LPECRS strain with an additional lysine-ON riboswitch for the control of lysE achieved a 21% increase in the yield of lysine compared to that of the C. glutamicum LPECRS strain and even a 89% increase in yield compared to that of the strain with deregulated aspartokinase. This work provides a useful approach to generate lysine-ON riboswitches for C. glutamicum metabolic engineering and demonstrates for the first time a synergetic effect of lysine-ON and -OFF riboswitches for improving lysine production in this industrially important microorganism. The approach can be used to dynamically control other genes and can be applied to other microorganisms.

Adição de lisina digestível em rações experimentais para juvenis de tilápia-do-nilo / Addition of lysine in experimental diets for Nile tilapia juveniles

Objetivou-se por meio deste estudo determinar a necessidade nutricional de lisina digestível em rações para juvenis de tilápia-do-nilo (Oreochromis niloticus). Setecentos e vinte peixes masculinizados (7,30±0,11g) foram alimentados durante 30 dias com oito rações (26,81% de proteína digestível e 3090kcal/kg de energia digestível da ração) contendo teores crescentes de lisina digestível (1,24; 1,36; 1,48; 1,60; 1,72; 1,84; 1,96 e 2,08%). As tilápias foram distribuídas em delineamento inteiramente ao acaso, com oito tratamentos, seis repetições e 15 peixes por unidade experimental. Foram avaliadas variáveis de desempenho (ganho de peso, taxa de crescimento específico, taxa de sobrevivência, consumo de ração, consumo de lisina digestível, conversão alimentar aparente, eficiência proteica para ganho, eficiência de lisina para ganho e eficiência de retenção de nitrogênio) e de composição corporal (teores de umidade, gordura, proteína, matéria mineral corporal e as taxas de deposição diária de proteína e gordura corporais). A elevação do teor de lisina digestível na ração não influenciou (P>0,05) o consumo de ração, a taxa de sobrevivência e os teores de umidade e de matéria mineral corporal, mas melhorou de forma quadrática (P<0,05) os demais parâmetros avaliados, com exceção do consumo de lisina e da eficiência de lisina para ganho, que aumentou e reduziu, respectivamente, de forma linear (P<0,05). Recomenda-se que rações para juvenis de tilápia-do-nilo devam conter 1,84% de lisina digestível para máximo ganho de peso...

The aim of this study was to determine the nutritional need of lysine in diets for juvenile Nile tilapia (Oreochromis niloticus). Seven hundred and twenty masculinized fish (7.30±0.11g) were fed for 30 days with eight diets (26.81% of digestible protein and 3090 kcal/kg digestible energy of feed) containing increasing levels of lysine (1.24, 1.36, 1.48, 1.60, 1.72, 1.84, 1.96 and 2.08%). The tilapia were distributed in a completely randomized design with eight treatments and six replicates of 15 fish per experimental unit. We evaluated the performance variables (weight gain, specific growth rate, survival rate, feed intake, digestible lysine intake, feed conversion, protein efficiency for gain, efficiency of lysine for gain and efficiency of retention nitrogen) and body composition (moisture, fat, protein, ash body and deposition rates of daily protein and fat). The high levels of dietary lysine did not affect (P>0.05) feed intake, the survival rate and the moisture and ash body, but improved (P<0.05) other parameters, except for lysine intake and efficiency of lysine for gain, which increased and decreased, respectively, linearly (P<0.05). It is recommended that diets for juvenile Nile tilapia should contain 1.84% digestible lysine for maximum weight gain...

Composición proximal y contenido de lípidos y colesterol de la carne de cerdos alimentados con harina de pijiguao (Bactris gasipaes Kunth) y lisina sintética

Se condujeron dos experimentos para evaluar la composición proximal y el contenido de lípidos y colesterol de la carne de cerdos alimentados con dietas en las que se añadió harina de pijiguao (HP) con y sin adición de lisina sintética (LIS). En el experimento I, se utilizaron 24 cerdos en crecimiento distribuidos al azar en seis tratamientos con tres niveles de HP (0, 16 y 32%) y dos niveles de LIS (0 y 0,27%). En el experimento II se utilizaron 16 cerdos en engorde alimentados con dos niveles de HP (0 y 17,50%) y dos niveles de LIS (0, y 0,27%). Al final de cada experimento (42 y 35 días, respectivamente), los cerdos fueron sacrificados y se obtuvieron muestras de lomo para determinar el contenido de proteína cruda, materia seca, humedad, cenizas, lípidos totales y colesterol. En el experimento I, las carnes provenientes de animales con 16% HP, presentaron más materia seca (26,45 g/100 g) y menos humedad (73,49 g/100 g) que las carnes de 32% HP (25,11 y 75,03 g/100g), respectivamente). Las carnes de los cerdos sin LIS presentaron mayor (p.

Two experiments were conducted to evaluate the proximal composition , lipids and cholesterol content of meat from pigs fed diets with peach-palm meal (PPM), with or without addition of synthetic lysine (LYS). In experiment I, 24 pigs were randomly allotted into six treatments with three levels of PPM (0, 16 and 32%) and two levels of LYS (0 and 0.27%). In experiment II, 16 finishing pigs were fed with two levels of PPM (0 and 17.50%) and two levels of LYS (0 and 0.27%). At the end of each experiment (42 and 35 d, respectively), pigs were slaughtered and loin samples were obtained to determine crude protein, dry matter, moisture, ash, total lipids, and cholesterol content. In experiment I, pork loin from 16% PPM had more dry matter (26.45 g/100 g) and less moisture (73.49 g/100g) than pork loin from 32% PPM (25.11 y 75.03 g/100g, respectively). Meat samples from pigs without LYS had higher (p.

Performance and carcass characteristics of swine at 50 kg fed diets with different energy levels and reduced levels of crude protein / Desempenho e características de carcaça de suínos aos 50 kg recebendo rações com diferentes níveis de energia e reduzido teor de proteína bruta

The aim of this work was to evaluate the effects of different levels of metabolic energy (ME) in diets with a reduced level of crude protein (CP) on the performance and carcass characteristics of growing swine. Thirty-two high- performance barrows (initial weight of 22.5 ± 1.43 kg and final weight of 50.81 ± 2.54 kg) were distributed in a randomized block design with four treatments (three levels of ME – 3,060,3,230 and 3,400 kcal/kg - in diets with 14% CP, and one control treatment with 3,230 kcal ME/kg and 18% CP) and four replicates. The diets were formulated with a true digestible basis of amino acids present in the feed, with methionine and threonine concentrations corrected as a function of lysine levels. The animals were slaughtered and their carcasses were studied. The ME levels did not affect the average weight gain, the feed:gain ratio, the backfat thickness or the loin eye area. A higher carcass yield and a higher carcass meat:fat ratio were observed in the animals that received the 3,230 and 3,060 kcal/kg ME levels. Compared to the control treatment, only the 3,400 kcal/kg ME level had a lower average daily feed intake. It is concluded that the CP level in diet should be reduced from 18% to 14% and the metabolizable energy level should be increased, thus reducing the feed intake and increasing the carcass meat:fat ratio without affecting the animal weight gain.

Objetivou-se, neste trabalho, avaliar diferentes níveis de energia metabolizável (EM) em rações com reduzido teor de proteína bruta (PB) através do desempenho e características de carcaça de suínos em crescimento. Foram utilizados 32 suínos machos castrados de elevado ganho de peso (peso inicial de 22,5 ± 1,43 kg e final de 50,81 ± 2,54 kg), distribuídos em delineamento de blocos casualizados, com quatro tratamentos (três níveis de EM - 3060, 3230 e 3400 kcal/kg - em rações com 14% PB e um tratamento controle com 3230 kcal EM/kg e 18% de PB) e quatro repetições. As rações foram formuladas com base na digestibilidade verdadeirados aminoácidos presentes nos alimentos, sendo a metionina e a treonina corrigidas em função dos níveis de lisina. Os animais foram abatidos e tiveram suas carcaças estudadas. Os níveis de EM não afetaram o ganho de peso médio diário, a conversão alimentar, a espessura de toucinho e a área de olho de lombo. Observou-se maior rendimento e relação carne:gordura na carcaça dos animais que receberam os níveis de 3230 e 3060 kcal EM, respectivamente. Comparado ao controle, apenas o nível de 3400 kcal/kg EM reduziu o consumo de ração médio diário. Conclui-se que o teor de PB da ração para suínos em crescimento pode ser reduzido de 18% para 14% e o nível de energia aumentado, propiciando menor consumo e maior relação carne: gordura sem afetar o ganho de peso dos animais.