RESUMEN
Aquaculture in Mexico has been developed by the cultivation of commercial species. In Tabasco, the cultivation of native species is mainly limited by the lack of nutrition studies to support its crop profitability. Among these species is the tropical gar (Atractosteus tropicus), which has great potential for cultivation. However, the nutritional value of carbohydrates in diets for this species which contribute to improved growth and survival, have not been evalulated,. Thus, in the present investigation, isoprotein and isolipid diets have been designed based on the substitution of cellulose by corn starch (D1: 0% starch-15% cellulose, D2: 7.5% starch-7.5% cellulose and D3: 15% starch-0% cellulose) and compared with a commercial trout diet (45% protein and 16% lipids). A total of 1800 larvae (0.008 ± 0.002 g and 10.5 ± LT 0.126 mm) were used, distributed in a recirculation system in order to evaluate growth and survival for 30 days. The results show higher growth and survival of 97% of larvae fed the D3 diet, while cannibalism in the species was mitigated. Major digestive enzyme activities occurred (acid protease, alkaline protease, trypsin, chymotrypsin, leucine aminopeptidase, carboxypeptidase A, lipase, α-glucosidase and amylase) for larvae fed D3. It is concluded that the contribution of corn starch (15%) replacing cellulose in the diet improves growth and survival of this species.
Asunto(s)
Acuicultura , Dieta/métodos , Peces/fisiología , Almidón , Amilasas/metabolismo , Animales , Leucil Aminopeptidasa/metabolismo , México , Péptido Hidrolasas/metabolismo , Tripsina/metabolismoRESUMEN
A study was performed in order to understand the development of digestive enzymes during initial ontogeny of Cichlasoma trimaculatum, for which the activity of acidic and alkaline proteases, lipases, amylases and phosphatases was determined by means of biochemical and electrophoretic analysis. Our results showed that the activity of alkaline proteases, trypsin and chymotrypsin is present from day 6 after hatching (dah) during exogenous feeding with Artemia nauplii. The activities of carboxypeptidase A and leucine aminopeptidase are present from the first days, increasing at 6 dah and reaching their maximum activity at 9 dah while acid protease activity started at 9 dah. Furthermore, the lipase activity is detected on 6 dah and keeps increasing and decreasing on 17 dah. Amylase activity is detected on 3 dah, presenting fluctuations until 45 dah, where it reaches its maximum activity. Acid and alkaline phosphatases are detected from 3 dah and reach a maximum activity between 13 and 19 dah. The SDS-PAGE electrophoresis revealed six types of bands in the alkaline proteases, with molecular weight between 113.4 and 20.4 kDa. First three bands appear on 6 dah, but it is until 11 dah when all isoforms appear. Based on these results, it is considered that this species completes its digestive enzymatic machinery from day 9 after hatching, therefore is recommended to perform the transition from live feed to inert feed at 15 dah.
Asunto(s)
Acuicultura/métodos , Cíclidos/crecimiento & desarrollo , Sistema Digestivo/enzimología , Regulación del Desarrollo de la Expresión Génica/fisiología , Regulación Enzimológica de la Expresión Génica/fisiología , Factores de Edad , Amilasas/metabolismo , Animales , Carboxipeptidasas A/metabolismo , Quimotripsina/metabolismo , Cíclidos/metabolismo , Sistema Digestivo/crecimiento & desarrollo , Electroforesis en Gel de Poliacrilamida , Leucil Aminopeptidasa/metabolismo , Lipasa/metabolismo , Péptido Hidrolasas/metabolismo , Tripsina/metabolismoRESUMEN
Enzyme activity was evaluated in the intestine of juvenile pacu, Piaractus mesopotamicus, fed diets containing 0, 10 or 20 % of lyophilized bovine colostrum (LBC) inclusion for either 30 or 60 days. The enzymes intestinal acid and alkaline phosphatase (ACP and ALP, respectively), nonspecific esterase (NSE), lipase (LIP), dipeptidyl aminopeptidase IV (DAP IV) and leucine aminopeptidase (LAP) were studied using histochemistry in four intestinal segments (S1, S2, S3 and rectum). Moderate activity of the DAP IV was detected in the three last intestinal segments, but no differences among the treatments were detected. Enzymes LAP, NSE and LIP were weakly stained in all intestinal segments and the inclusion of 10 or 20 % of LBC in the diet commanded a moderate reaction to NSE in the S3 segment at day 60. ACP activity was detected only in the brush border of the S1 segment of fish fed 0 % LBC for either 30 or 60 days. The activity of ALP was very strong in the first intestinal segment, but a weak reaction was seen in the last segments. The inclusion of 20 % of LBC changed the pattern of staining to the ALP, eliciting moderate staining in S2 at day 30 and S1 at day 60. The consumption of diets containing LBC by juvenile pacu did not have significant implications in intestinal enzymatic activity, which still was not fully stimulated.
Asunto(s)
Characiformes/metabolismo , Dieta , Intestinos/enzimología , Fosfatasa Ácida/metabolismo , Fosfatasa Alcalina/metabolismo , Animales , Carboxilesterasa/metabolismo , Bovinos , Calostro/química , Liofilización , Técnicas Histológicas/veterinaria , Leucil Aminopeptidasa/metabolismo , Lipasa/metabolismo , Cloruro de TolonioRESUMEN
Tropical gar (Atractosteus tropicus) is an economically and socially important freshwater species from Southeastern Mexico, with a high aquaculture potential. With this in mind, the purpose of this study was to characterize the digestive proteases of tropical gar juveniles through biochemical and electrophoretic analyses. Twenty specimens with an average weight of 73.6 ± 12.7 g were used to obtain stomach and intestinal tissue from which multienzymatic extracts were prepared. The general activities of the acid and alkaline proteases were evaluated, as well as the specific activities of trypsin, chymotrypsin, leucine aminopeptidase and carboxypeptidase A. The effect of the pH and temperature on the proteases was also analyzed, together with the composition of the multienzymatic extracts using protease inhibitors and electrophoretic tests. Results showed that A. tropicus have a functional stomach in which protein hydrolysis starts with pepsin and which contains endo- and exopeptidases (trypsin, chymotrypsin, leucine aminopeptidase and carboxypeptidase A) and proteases that are resistant to high temperatures (45 and 55 °C for alkaline and acid proteases, respectively) and pH values. Using zymogram technique, we found two acid protease isoforms (0.35 and 0.71 rf) and five alkaline protease isoforms (83.7, 43.7, 27.5, 24.0 and 19.4 kDa), which decrease or disappear with the different inhibitors. Thus, this species is considered to be a carnivore capable of adapting to its environment by consuming different types of proteins from preys and also could adapt rapidly to consume a compound diet with different animal protein sources.
Asunto(s)
Digestión/fisiología , Peces/metabolismo , Tracto Gastrointestinal/enzimología , Péptido Hidrolasas/metabolismo , Animales , Acuicultura/métodos , Carboxipeptidasas A/metabolismo , Quimotripsina/metabolismo , Electroforesis/veterinaria , Peces/fisiología , Concentración de Iones de Hidrógeno , Isoenzimas/metabolismo , Leucil Aminopeptidasa/metabolismo , México , Estadísticas no Paramétricas , Temperatura , Tripsina/metabolismoRESUMEN
Leucine aminopeptidase from Vibrio proteolyticus is a broad specificity N-terminal aminopeptidase that is widely used in pharmaceutical processes where the removal of N-terminal residues in recombinant proteins is required. We previously reported the expression of a heterologous construction of the mature protein fused to a 6-histidine tag that presents a reasonable refolding rate for its use at industrial level. Here, we investigate this recombinant leucine aminopeptidase (rLAP) to explain the gain of activity observed when incubated at 37 °C after its production. Unfolding transitions of rLAP as a function of urea concentration were monitored by circular dichroism (CD) and fluorescence (FL) spectroscopy exhibiting single transitions by both techniques. Free energy change for unfolding measured by CD and FL spectroscopy are 2.8 ± 0.4 and 3.7 ± 0.4 kcal mol(-1), respectively. Thermal stability conformation of rLAP is 2.6 ± 0.1 and 6.1 kcal mol(-1) for CD and Nano-Differential Scanning Calorimetry (Nano-DSC), respectively. Enzyme activity was assessed with L-leucine-p-nitroanilide (L-pNA) as substrate. The catalytic efficiency was 3.87 ± 0.10 min(-1) µM(-1) at 37 °C and pH 8.0. Kinetic and conformation studies show differences between the enzyme native and rLAP; however rLAP is selective and specific to remove N-terminal groups from amino acids.
Asunto(s)
Leucil Aminopeptidasa/química , Proteínas Recombinantes/química , Activación Enzimática , Estabilidad de Enzimas , Cinética , Leucil Aminopeptidasa/metabolismo , Conformación Proteica , Replegamiento Proteico/efectos de los fármacos , Desplegamiento Proteico/efectos de los fármacos , Proteínas Recombinantes/metabolismo , Termodinámica , Urea/farmacología , Vibrio/enzimologíaRESUMEN
Realizou-se um levantamento bibliográfico, com artigos de revisão, analisando e discutindo os trabalhos publicados sobre os efeitos da leucina aminopeptidase e aminopeptidase A no trabalho de parto pré-termo e na pré-eclampsia. A proposta deste trabalho sobre o tema é que grande parte das questões de saúde materna parece pueril, principalmente quanto ao atendimento voltado para os cuidados maternos, no qual, a cada 20minutos, morre uma mulher em decorrência de parto, no mundo todo. Por isso, tais doenças poderão receber mais atenção do que outras. Esse fato fez com que houvesse certa preocupação com o índice de natalidade e morbidade materna, bem como morbidade e mortalidade perinatal. Portanto, abordou-se sobre sua biologia geral, fisiologia de reprodução, síntese de evolução genética, habitação, alimentação, manejo, dor e eutanásia, técnicas de riscos desenvolvidos na experimentação animal, estudos de experimentos farmacológicos e toxicológicos observados dentro dos artigos de revisão. Embora tendências atuais preconizem a utilização de métodos alternativos (estudo in vitro), os modelos animais, como as ratas, apresentam como principal vantagem o fornecimento de informações sobre o organismo como um todo, fato que não é obtido com outros métodos, o que ainda possibilita o seu emprego em pesquisas científicas.
We have carried out a literature review, with review articles, analyzing and discussing the works that have already been published on the effects of leucine aminopeptidade and aminopeptidase A in pre-term labor and preeclampsia. The proposal of this work on the subject is that most of the issues of maternal health seems childish, especially for service oriented maternity care, where, every 20 minutes, a woman dies due to childbirth, worldwide. Therefore, such diseases may receive more attention than others. This led to worry about the birth rate and maternal morbidity and perinatal morbidity and mortality. Therefore, it was addressed their general biology, physiology of reproduction, synthesis of evolution, genetics, housing, feeding, pain and euthanasia techniques developed for animal experimentation risks, studies of pharmacological and toxicological experiments observed within the review articles. Although current trends have preconized the use of alternative methods (in vitro study), animal models, such as rats, have as main advantage the provision of information on the organism as a whole, a fact that is not achieved with other methods, which also allows its use in scientific research.
Asunto(s)
Animales , Ratas , Cistinil Aminopeptidasa/metabolismo , Preeclampsia/enzimología , Preeclampsia/etiología , Trabajo de Parto Prematuro/enzimología , Cistinil Aminopeptidasa/sangre , Inmunohistoquímica/métodos , Leucil Aminopeptidasa/metabolismo , Leucil Aminopeptidasa/sangre , Ratas Wistar , Sulfato de Magnesio/efectos adversos , Trabajo de Parto Prematuro/sangreRESUMEN
BACKGROUND: Pathogens depend on peptidase activities to accomplish many physiological processes, including interaction with their hosts, highlighting parasitic peptidases as potential drug targets. In this study, a major leucyl aminopeptidolytic activity was identified in Trypanosoma cruzi, the aetiological agent of Chagas disease. RESULTS: The enzyme was isolated from epimastigote forms of the parasite by a two-step chromatographic procedure and associated with a single 330-kDa homohexameric protein as determined by sedimentation velocity and light scattering experiments. Peptide mass fingerprinting identified the enzyme as the predicted T. cruzi aminopeptidase EAN97960. Molecular and enzymatic analysis indicated that this leucyl aminopeptidase of T. cruzi (LAPTc) belongs to the peptidase family M17 or leucyl aminopeptidase family. LAPTc has a strong dependence on neutral pH, is mesophilic and retains its oligomeric form up to 80°C. Conversely, its recombinant form is thermophilic and requires alkaline pH. CONCLUSIONS: LAPTc is a 330-kDa homohexameric metalloaminopeptidase expressed by all T. cruzi forms and mediates the major parasite leucyl aminopeptidolytic activity. Since biosynthetic pathways for essential amino acids, including leucine, are lacking in T. cruzi, LAPTc could have a function in nutritional supply.
Asunto(s)
Leucil Aminopeptidasa/química , Leucil Aminopeptidasa/metabolismo , Multimerización de Proteína , Trypanosoma cruzi/enzimología , Secuencia de Aminoácidos , Citoplasma/metabolismo , Descubrimiento de Drogas , Hidrólisis , Leucil Aminopeptidasa/clasificación , Leucil Aminopeptidasa/aislamiento & purificación , Datos de Secuencia Molecular , Filogenia , Estructura Cuaternaria de Proteína , Transporte de Proteínas , Alineación de Secuencia , Trypanosoma cruzi/citología , Trypanosoma cruzi/efectos de los fármacosRESUMEN
A new leucyl aminopeptidase activity has been identified in the fission yeast Schizosaccharomyces pombe. The enzyme, which has been purified and named leucyl aminopeptidase yspII (LAP yspII), had a molecular mass of 320 and 54 kDa by gel filtration and SDS/PAGE, respectively, suggesting a homohexameric structure. The enzyme cleaved synthetic aminoacyl-4-nitroanilides at an optimum of pH 8.5, and preferred leucine and methionine as N-terminal amino acids. A clear dependence on Mn2+ concentration for activity was found, and an apparent association constant of 0.33 mM was calculated for the metal ion. Bestatin behaved as a competitive inhibitor of LAP yspII (K(i) = 0.14 microM), while chelating agents such as chloroquine, EDTA and 1,10-phenanthroline also reduced enzyme activity. A MALDI-MS analysis, followed by sequencing of two of the resulting peptides, showed that LAP yspII undoubtedly corresponds to the putative aminopeptidase C13A11.05 identified in the S. pombe genome project. The protein exhibited nearly 40% sequence identity to fungal and mammalian aminopeptidases belonging to the M17 family of metallopeptidases. Catalytic residues (Lys292 and Arg366), as well as those involved in coordination with the cocatalytic metal ions (Lys280, Asp285, Asp303, Asp362 and Glu364) and those forming the hydrophobic pocket for substrate binding (Met300, Asn360, Ala363, Thr390, Leu391, Ala483 and Met486), were perfectly conserved among all known aminopeptidases. The S. pombe enzyme is predicted to be formed two clearly distinguished domains with a well conserved C-terminal catalytic domain showing a characteristic topology of eight beta-sheets surrounded by alpha-helical segments in the form of a saddle.
Asunto(s)
Citosol/enzimología , Leucil Aminopeptidasa/química , Leucil Aminopeptidasa/metabolismo , Schizosaccharomyces/enzimología , Secuencia de Aminoácidos , Secuencia de Bases , Sitios de Unión , Dominio Catalítico , Cationes Bivalentes/metabolismo , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Concentración de Iones de Hidrógeno , Hidrólisis , Cinética , Leucina/análogos & derivados , Leucina/farmacología , Leucil Aminopeptidasa/antagonistas & inhibidores , Leucil Aminopeptidasa/genética , Leucil Aminopeptidasa/aislamiento & purificación , Manganeso/metabolismo , Modelos Químicos , Modelos Moleculares , Datos de Secuencia Molecular , Peso Molecular , Unión Proteica , Conformación Proteica , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Homología de Secuencia de Aminoácido , Especificidad por SustratoRESUMEN
Changes in the expression of genes were observed during development in populations of Anopheles (Anopheles) intermedius and Anopheles (Anopheles) mattogrossensis. Esterase showed seven zones of activity: EST1 was present in all developmental stages of both species; EST2 was observed only in larvae of A. intermedius and larvae and pupae of A. mattogrossensis, with greater activity in pupae; EST3 and EST5 were present in all developmental stages, with greater intensity in larvae; EST4 and EST6 showed weak activity in larvae of A. mattogrossensis and was not found in A. intermedius. Leucine aminopeptidase showed four zones of activity, of which LAP1 and LAP2 were found in all stages of A. intermedius, with highest activity in larvae, and in larvae only of A. mattogrossensis. LAP3 was detected in all stages of A. mattogrossensis and in larvae only of A. intermedius. LAP4 was detected only in larvae and pupae of A. mattogrossensis, with greater intensity in pupae. alpha-Glycerophosphate dehydrogenase showed a single zone of activity, detected in older fourth-instar larvae and becoming more intense from the pupal stage onwards.
Asunto(s)
Anopheles/genética , Variación Genética , Animales , Anopheles/clasificación , Anopheles/enzimología , Brasil , Electroforesis , Esterasas/genética , Esterasas/metabolismo , Regulación Enzimológica de la Expresión Génica , Glicerolfosfato Deshidrogenasa/genética , Glicerolfosfato Deshidrogenasa/metabolismo , Leucil Aminopeptidasa/genética , Leucil Aminopeptidasa/metabolismoRESUMEN
Changes in the expression of genes were observed during development in populations of Anopheles (Anopheles) intermedius and Anopheles (Anopheles) mattogrossensis. Esterase showed seven zones of activity: EST1 was present in all developmental stages of both species; EST2 was observed only in larvae of A. intermedius and larvae and pupae of A. mattogrossensis, with greater activity in pupae; EST3 and EST5 were present in all development stages, with greater intensity in larvae; EST4 and EST6 showed weak activity in larvae of A. mattogrossensis and was not found in A. intermedius. Leucine aminopeptidase showed four zones of activity, of which LAP1 and LAP2 were found in all stages of A. intermedius, with highest activity in larvae, and in only of A. mattogrossensis. LAP3 was detected in all stages of A. mattogrossensis and in larvae only of A. intermedius. LAP4 was detected only in larvae and pupae of A. mattogrossensis, with greater intensity in pupae. Alpha-Glycerophosphate dehydrogenase showed a single zone of activity, detected in older fourth-instar larvae and becoming more intense from the pupal stage onwards.
Asunto(s)
Animales , Anopheles/genética , Variación Genética , Anopheles/enzimología , Brasil , Electroforesis , Esterasas/genética , Esterasas/metabolismo , Regulación Enzimológica de la Expresión Génica , Glicerolfosfato Deshidrogenasa/genética , Glicerolfosfato Deshidrogenasa/metabolismo , Leucil Aminopeptidasa/genética , Leucil Aminopeptidasa/metabolismoRESUMEN
Leucyl aminopeptidase (LAP; EC 3.4.11.1) activity was purified from crude extracts of the marine unicellular algae Gonyaulax polyedra by a combination of hydrophobic interaction with phenyl sepharose, DEAE-cellulose, and mono-Q HR5/5 ion-exchange chromatography. The undenaturated protein has a molecular mass of about 110 kD and based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the enzyme appears to be composed of two possibly identical subunits of 55 kD. The identity of the protein was confirmed by a cross-reaction of the purified protein with an antibody raised against a commercial LAP. Biochemical characterization showed that the Gonyaulax enzyme was similar to most of the previously described LAPs. Gonyaulax LAP is a metallo-enzyme since EDTA and 1,10-phenathroline significantly inhibited activity. Addition of the metal ions Zn(2+), Cu(2+) inhibited 80% of LAP activity, suggesting they are not the natural cofactors of the enzyme. Other metals, such as Ca(2+), Co(2+), Mn(2+), or Mg(2+) (concentrations up to 4 mM), caused no alteration in the total activity of Gonyaulax LAP.
Asunto(s)
Dinoflagelados/enzimología , Leucil Aminopeptidasa/aislamiento & purificación , Metaloproteínas/aislamiento & purificación , Animales , Cromatografía/métodos , Ritmo Circadiano , Reacciones Cruzadas , Leucil Aminopeptidasa/inmunología , Leucil Aminopeptidasa/metabolismo , Metaloproteínas/inmunología , Metaloproteínas/metabolismo , Proteínas/metabolismo , Ubiquitinas/metabolismoRESUMEN
An aminopeptidase activity capable of hydrolyzing different aminomethylcoumarin amino acids, but mainly leucine-7-amino-4-methylcoumarin (Leu-NHMc), was detected in deoxycholic soluble extracts from adult Fasciola hepatica. The enzyme (EC 3.4.11.1) was partially purified by gel filtration and EAH-Sepharose affinity chromatography using bestatin as a ligand. Results obtained by gel filtration, direct fluorogenic substrate analysis in polyacrylamide gel, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis suggest that in a native form the enzyme might be aggregated as a high molecular weight complex. By affinity chromatography on concanavalin A Sepharose, the enzyme did not bond to the column showing that it lacks mannose residues. The F. hepatica aminopeptidase was characterized as a metalloproteinase based on its activation by Mn2+ and Mg2+, and its inhibition by EDTA, 1,10-phenanthroline, and bestatin. It has an optimal activity at a pH between 8.0 and 8.5. Histochemical localization revealed strong leucine naphthylamidase activity at the cells lining the gut epithelium of the parasite.
Asunto(s)
Fasciola hepatica/enzimología , Leucil Aminopeptidasa/metabolismo , Animales , Bovinos , Cromatografía de Afinidad , Cromatografía en Gel , Cumarinas/metabolismo , Ácido Desoxicólico/metabolismo , Electroforesis en Gel de Poliacrilamida , Histocitoquímica , Concentración de Iones de Hidrógeno , Hidrólisis , Indicadores y Reactivos/metabolismo , Leucil Aminopeptidasa/antagonistas & inhibidores , Leucil Aminopeptidasa/química , Leucil Aminopeptidasa/aislamiento & purificación , Metales/farmacología , Peso Molecular , Inhibidores de Proteasas/farmacología , Especificidad por SustratoRESUMEN
The esterases, leucine aminopeptidase and alpha-glycerophosphate dehydrogenase revealed modifications in gene expressions during the development of Anopheles darlingi. The esterases showed five activity bands, 1 and 2 being more deeply stained during the larval stages than in pupae or adults, esterases 3 and 4 more deeply stained in pupae and adults whereas esterase 5 was present throughout development. Leucine aminopeptidase showed five activity bands: LAP2 and LAP5 were characteristic of larvae, LAP3 was specific for pupae and adults, LAP4 was detected only in pupae, and LAP1 and LAP6 were detected in all stages. alpha-Glycerophosphate dehydrogenase presented one activity band on starch gel whose intensity increased with development. Two activity bands were detected on polyacrylamide gel (alpha-GPDH1 and alpha-GPDH2) in 4th-instar larvae (old pigmented larvae) and this activity increased with development.
Asunto(s)
Anopheles/genética , Esterasas/genética , Glicerolfosfato Deshidrogenasa/genética , Leucil Aminopeptidasa/genética , Animales , Anopheles/enzimología , Anopheles/crecimiento & desarrollo , Esterasas/metabolismo , Expresión Génica , Glicerolfosfato Deshidrogenasa/metabolismo , Leucil Aminopeptidasa/metabolismoRESUMEN
The esterases, leucine aminopeptidase and alpha-glycerophosphate dehydrogenase revealed modifications in gene expressions during the development of Anopheles darlingi. The esterases showed five activity bands, 1 and 2 being more deeply stained during the larval stages than in pupae or adults, esterases 3 and 4 more deeply stained in pupae and adults whereas esterase 5 was present throughout development. Leucine aminopeptidase showed five activity bands: LAP2 and LAP5 were characteristic of larvae, LAP3 was specific for pupae and adults, LAP4 was detected only in pupae, and LAP1 and LAP6 were detected in all stages. Alpha-Glycerophosphate dehydrogenase presented one activity band on starch gel whose intensity increased with development. Two activity bands were detected on polyacrylamide gel (alpha-GPDH1 and alpha-GPDH2) in 4th-instar larvae (old pigmented larvae) and this activity increased with development.
Asunto(s)
Animales , Anopheles/genética , Esterasas/genética , Expresión Génica , Variación Genética , Glicerolfosfato Deshidrogenasa/genética , Leucil Aminopeptidasa/genética , Anopheles/enzimología , Anopheles/crecimiento & desarrollo , Electroforesis en Gel de Poliacrilamida , Electroforesis en Gel de Almidón , Esterasas/metabolismo , Glicerolfosfato Deshidrogenasa/metabolismo , Isoenzimas/genética , Isoenzimas/metabolismo , Leucil Aminopeptidasa/metabolismoRESUMEN
The effect of substrate phosphorylation on the susceptibility to exopeptidasic attack by leucyl aminopeptidase of swine kidney, alanyl aminopeptidase from human liver and aminopeptidase N of Escherichia coli was investigated using a synthetic heptapeptide (L-R-R-A-S-L-G) and its phosphorylated derivative. The enzyme-catalyzed products were analyzed by thin layer chromatography and electrophoresis. The sensitivities of peptide and phosphopeptide to leucyl aminopeptidase digestion were then compared. Data obtained indicated that when phosphopeptide was used as substrate one main product accumulated, which corresponded to the fragment A-S(P)-L-G, while unphosphorylated peptide was completely degraded to its constituent amino acids. Identical results were obtained using aminopeptidase N of E. coli. Using alanyl aminopeptidase as enzyme, the results obtained were essentially similar, since the exopeptidasic activity on the phosphorylated peptide was strongly hampered in the vicinity of phosphoseryl residue leading to accumulation of the same phosphorylated product, although this enzyme could not completely degrade the unphosphorylated peptide. It was concluded that phosphorylation of substrates does effect enzymic degradation of proteins.
Asunto(s)
Antígenos CD13/metabolismo , Leucil Aminopeptidasa/metabolismo , Oligopéptidos/metabolismo , Secuencia de Aminoácidos , Animales , Escherichia coli/enzimología , Humanos , Hidrólisis , Datos de Secuencia Molecular , Fosforilación , Especificidad por Sustrato , PorcinosRESUMEN
The interaction between malnutrition and exposure to a mucosal damaging agent, difluoromethylornithine (DFMO), was examined by monitoring the small-intestinal changes in weanling rats. Malnutrition as induced by the expanded-litter method resulted in severe reduction in body weights in the expanded litters as compared to normal litters. Subsequent treatment of malnourished and well-nourished pups with DFMO for 7 days resulted in decreases in small-intestinal weights and enzyme contents. A 2 factors (well-nourished and malnourished) by 2 factors (DFMO-treated and nontreated) analysis of variance showed no interaction between malnutrition and DFMO treatment in terms of food intake, total mucosal protein, and contents of enterokinase, leucine aminopeptidase and sucrase. Very slight and insignificant interactions (p less than or equal to 0.2) were found for body weights, intestinal weights and total DNA content. Only one parameter studied, the maltase content, showed significant interaction between malnutrition and DFMO treatment (p less than 0.05). Three weeks after the withdrawal of DFMO, essentially all the changes caused by DFMO recovered. But those changes caused by malnutrition did not, such that the malnourished group, whether treated with DFMO or not, still remained significantly less than the control group in their small-intestinal parameters. Analysis of variance showed no interaction between malnutrition and DFMO treatment in the recovery phase. The results suggest that malnutrition is a more important factor in determining the intestinal damage and that malnutrition in the immediate postnatal period does not increase the sensitivity of the small intestine to the damaging effect of DFMO.
Asunto(s)
Eflornitina/toxicidad , Mucosa Intestinal/efectos de los fármacos , Trastornos Nutricionales/complicaciones , Animales , Animales Lactantes , División Celular/efectos de los fármacos , Femenino , Glicósido Hidrolasas/metabolismo , Mucosa Intestinal/enzimología , Mucosa Intestinal/patología , Leucil Aminopeptidasa/metabolismo , Masculino , Trastornos Nutricionales/enzimología , Fosfotransferasas/metabolismo , Ratas , Ratas EndogámicasRESUMEN
Se estudia el efecto que produce el acetato de hidrocortisona y la prednisolona en ratas de 12 días de nacidas que habían sido previamente inyectadas durante 3 días consecutivos con una dosis de 50 mg/kg de peso. Se observó que el efecto del acetato de hidrocortisona fue más intenso que el de la prednisolona al incrementar la actividad de la sacarasa y la leucina aminopeptidasa; y que el efecto de la prednisolona fue más intenso que el del acetato de hidrocortisona al incrementar la actividad de la fosfatasa alcalina y al disminuir la ß galactosidasa ácida y la catepsina D. Además, ninguna de las dos hormonas provocó cambio en la lactato deshidrogenasa. Se concluye que aunque el efecto de ambas hormonas es diferente desde el punto de vista cuantitativo, la prednisolona es capaz de producir los cambios de los niveles enzimáticos de forma similar a los que provoca el acetato de hidrocortisona
Asunto(s)
Ratas , Animales , Estudio Comparativo , Acetatos , Hidrocortisona , Prednisolona , Sacarasa/metabolismo , beta-Galactosidasa/metabolismo , Catepsina D/metabolismo , Fosfatasa Alcalina/metabolismo , Leucil Aminopeptidasa/metabolismo , L-Lactato Deshidrogenasa/metabolismo , Intestino Delgado/enzimologíaRESUMEN
Se estudia el efecto que produce el acetato de hidrocortisona y la prednisolona en ratas de 12 días de nacidas que habían sido previamente inyectadas durante 3 días consecutivos con una dosis de 50 mg/kg de peso. Se observó que el efecto del acetato de hidrocortisona fue más intenso que el de la prednisolona al incrementar la actividad de la sacarasa y la leucina aminopeptidasa; y que el efecto de la prednisolona fue más intenso que el del acetato de hidrocortisona al incrementar la actividad de la fosfatasa alcalina y al disminuir la ß galactosidasa ácida y la catepsina D. Además, ninguna de las dos hormonas provocó cambio en la lactato deshidrogenasa. Se concluye que aunque el efecto de ambas hormonas es diferente desde el punto de vista cuantitativo, la prednisolona es capaz de producir los cambios de los niveles enzimáticos de forma similar a los que provoca el acetato de hidrocortisona
Asunto(s)
Ratas , Animales , Acetatos , Fosfatasa Alcalina/metabolismo , beta-Galactosidasa/metabolismo , Catepsina D/metabolismo , Hidrocortisona , Intestino Delgado/enzimología , L-Lactato Deshidrogenasa/metabolismo , Leucil Aminopeptidasa/metabolismo , Prednisolona , Sacarasa/metabolismoRESUMEN
Isozymes of leucine aminopeptidase (LAP) in leaf tissue of the cultivated chenopods (Chenopodium quinoa and C. nuttalliae) and their sympatric weedy relatives (C. hircinum and C. berlandieri) can be electrophoretically resolved into a sum total of five anodally migrating bands. These are the products of two unlinked gene loci which apparently reflect genetic duplication via allotetraploidy. Accessions from the Andean and Mexican areas of cultivation are characterized by a lack of electrophoretically detectable variation. Andean weed and cultigen accessions appear to be genetically identical at both Lap loci, as do weed and cultigen material from Mexico. The two cultigens, and their sympatric weeds, can be differentiated by variation at the Lap-B locus, whereas they are monomorphic at Lap-A. This locus is, however, highly polymorphic in weedy C. berlandieri populations of western North America.