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1.
J Dairy Sci ; 95(11): 6665-76, 2012 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-22959939

RESUMEN

The objective of this study was to evaluate transition cow performance when brown midrib corn silage (BMRCS; Mycogen F2F444) was included in the diet during the transition period, and to determine if any production response occurring during the first 3 wk of lactation would persist from wk 4 to 15 when a common diet was fed. Seventy Holstein dairy cows were blocked by parity (either second or third and greater) and calving date and randomly assigned to the CCS (a mixture of varieties of conventional corn silage) or BMRCS treatment. Diets were formulated with the objective of keeping all ration parameters the same, with the exception of neutral detergent fiber digestibility. Neutral detergent fiber digestibility values (30 h) for CCS and BMRCS averaged 56.8 and 73.8%, respectively. Prepartum rations contained 47% corn silage, 18% wheat straw, 7% alfalfa haylage, and 28% concentrate, and averaged 45% neutral detergent fiber (DM basis). Postpartum rations contained 40% corn silage, 15% alfalfa haylage, 1% straw, and 44% concentrate. Milk weights (3×/d) and dry matter intake were recorded daily, and milk composition was measured weekly. Cows fed BMRCS had higher dry matter intake during the 2-wk period before calving (14.3 vs. 13.2 kg/d) and the 3-wk period after calving (20.1 vs. 18.1 kg/d) than did cows fed CCS. Yields of milk, solids, and lactose were increased, whereas a trend was observed for a reduction in somatic cell counts and linear scores in the postpartum period for cows receiving BMRCS during the transition. A significant carryover effect of BMRCS was observed on production from wk 4 to 15 when the common diet was fed, with yields of protein (1.36 vs. 1.30 kg/d), lactose (2.24 vs. 2.12 kg/d), and solids (5.82 vs. 5.51 kg/d) increasing significantly, and yields of fat-corrected milk, energy-corrected milk, and fat tending to increase during this period for cows that had been fed BMRCS. The increased intakes during the last 2 wk of the prepartum period in the BMRCS treatment were likely because of a reduction in fill, whereas the increased intakes in the postpartum period in cows fed the BMRCS were either because of the higher intakes during the prepartum period or because of a reduction in fill limitations in the postpartum period. The carryover response in wk 4 to 15 may have resulted from cows that received BMRCS during the transition period being in a more positive nutrient balance than cows fed CCS. The results of this study indicate the importance that digestible NDF can have in transition diets and the long-term production responses that can occur when intake is increased in the transition period.


Asunto(s)
Ingestión de Alimentos/fisiología , Leche/química , Ensilaje , Zea mays , Ácido 3-Hidroxibutírico/análisis , Fenómenos Fisiológicos Nutricionales de los Animales/fisiología , Animales , Bovinos , Dieta/veterinaria , Metabolismo Energético/fisiología , Ácidos Grasos no Esterificados/análisis , Femenino , Lactancia/fisiología , Paridad , Periodo Periparto/fisiología , Embarazo , Ensilaje/análisis , Zea mays/química
2.
Science ; 246(4927): 235-40, 1989 Oct 13.
Artículo en Inglés | MEDLINE | ID: mdl-2552582

RESUMEN

The beta-adrenergic receptor kinase (beta-ARK), which specifically phosphorylates only the agonist-occupied form of the beta-adrenergic and closely related receptors, appears to be important in mediating rapid agonist-specific (homologous) desensitization. The structure of this enzyme was elucidated by isolating clones from a bovine brain complementary DNA library through the use of oligonucleotide probes derived from partial amino acid sequence. The beta-ARK cDNA codes for a protein of 689 amino acids (79.7 kilodaltons) with a protein kinase catalytic domain that bears greatest sequence similarity to protein kinase C and the cyclic adenosine monophosphate (cyclic AMP)--dependent protein kinase. When this clone was inserted into a mammalian expression vector and transfected into COS-7 cells, a protein that specifically phosphorylated the agonist-occupied form of the beta 2-adrenergic receptor and phosphorylated, much more weakly, the light-bleached form of rhodopsin was expressed. RNA blot analysis revealed a messenger RNA of four kilobases with highest amounts in brain and spleen. Genomic DNA blot analysis also suggests that beta-ARK may be the first sequenced member of a multigene family of receptor kinases.


Asunto(s)
Proteínas Quinasas Dependientes de AMP Cíclico , Familia de Multigenes/genética , Proteínas Quinasas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Bovinos , Clonación Molecular , Datos de Secuencia Molecular , Especificidad de Órganos , Fosforilación , Proteínas Quinasas/biosíntesis , Proteínas Quinasas/fisiología , Receptores Adrenérgicos beta/metabolismo , Homología de Secuencia de Ácido Nucleico , Especificidad por Sustrato , Quinasas de Receptores Adrenérgicos beta
3.
FEBS Lett ; 283(1): 122-6, 1991 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-2037065

RESUMEN

The beta-adrenergic receptor kinase (beta ARK) mediates agonist-dependent phosphorylation of the beta 2-adrenergic and related G protein-coupled receptors. A cDNA encoding bovine beta ARK has recently been isolated. In this work we have isolated a cDNA encoding human beta ARK from a retinal cDNA library. The cDNA encodes a protein of 689 amino acids with an overall 98.0% amino acid and 92.5% nucleotide identity with bovine beta ARK. Chromosomal location of the human beta ARK gene was determined by correlating the presence of the beta ARK locus with retention of a specific human chromosome in a rodent-human hybrid panel. This analysis revealed that the human beta ARK locus segregated with the long arm of chromosome 11, centromeric to 11q13.


Asunto(s)
Mapeo Cromosómico , Proteínas Quinasas Dependientes de AMP Cíclico , ADN/genética , Proteínas Quinasas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Southern Blotting , Bandeo Cromosómico , Cromosomas Humanos Par 11 , Clonación Molecular , Quinasa 2 del Receptor Acoplado a Proteína-G , Humanos , Células Híbridas , Datos de Secuencia Molecular , Roedores , Quinasas de Receptores Adrenérgicos beta
4.
J Anim Sci ; 73(1): 267-77, 1995 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-7601743

RESUMEN

Accurate prediction of forage biological values and performance with animals fed forages requires accurately accounting for factors that influence animal requirements and feedstuff utilization. The Cornell Net Carbohydrate and Protein System (CNCPS) is an application model that uses a combination of mechanistic and empirical approaches to account for the effects of variation in animal factors and feed carbohydrate and protein fractions on animal performance. Thus, accurate animal and environmental descriptions, DMI, feed carbohydrate, and protein fractions and their digestion rates are required inputs. In 25 growth periods with calves fed high-forage diets, the CNCPS accounted for 74, 81, and 83%, respectively, of the variation in ADG predicted to be supported by the ME, metabolizable protein, and essential amino acid intake, the first-limiting of all three accounting for 81% of the variation with a -1% bias. Thus, the CNCPS can be used to accurately describe forage quality and the effects of changes in forage composition on animal performance. The model was sensitive to variations in NDF, CP, protein solubility, NDF and starch digestion rates, feed and microbial amino acid composition, maintenance protein requirement, body protein amino acid content, and the coefficient of efficiency of use of absorbed protein. Analysis of several trials indicates an improved efficiency of ME use with improved amino acid balances. Uses of the CNCPS discussed include interpreting, planning and applying research, teaching, developing tables of requirements and biological values for feeds, complex nutritional accounting, and predicting performance and profits.


Asunto(s)
Alimentación Animal/normas , Bovinos/crecimiento & desarrollo , Carbohidratos de la Dieta/normas , Proteínas en la Dieta/normas , Modelos Biológicos , Alimentación Animal/análisis , Animales , Carbohidratos de la Dieta/análisis , Fibras de la Dieta , Proteínas en la Dieta/análisis
5.
Am J Vet Res ; 53(11): 2153-6, 1992 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-1466514

RESUMEN

Intestinal ischemia was induced and maintained for 60 minutes in male Sprague-Dawley rats weighing 175 to 225 g. Prior to reperfusion, the following drugs were administered via the caudal vena cava: 0.9% NaCl (0.5 ml), superoxide dismutase (SOD; 1,000 IU/kg of body weight), polyethylene glycol-conjugated SOD (PEG-SOD; 1,000 IU/kg), or the 21-aminosteroids, U74006F (3 mg/kg) or U78715G (3 mg/kg). A sham-operated control group was included. Animals from each group were euthanatized at 5 periods of reperfusion: 5 minutes, 30 minutes, 18 hours, 3 days, and 7 days after reperfusion. Fixed tissues were embedded in paraffin, sectioned at 5 microns, and stained with H&E. Villi profiled in cross section were measured from the crypt villus junction to the tip of the villus. The mean villus height for each rat was calculated and compared by two-way ANOVA to determine the effects of time and treatment. Villus height was maintained after 30 minutes of reperfusion in rats of the sham- and U74006F-treated groups; U78715G and SOD treatment attenuated the loss in villus height, and villus height was not maintained in the PEG-SOD- and 0.9% NaCl-treated rats. In all rats, villus height was comparable to, or was greater than villus height in sham-operated controls by 18 hours after reperfusion in all animals and remained constant through 7 days. Administration of the 21-aminosteroids maintained villus height after ischemia and reperfusion. Treatment with PEG-SOD did not maintain villus height to the degree observed in rats treated with SOD.


Asunto(s)
Depuradores de Radicales Libres , Mucosa Intestinal/efectos de los fármacos , Peróxidos Lipídicos/antagonistas & inhibidores , Daño por Reperfusión/patología , Animales , Estudios de Evaluación como Asunto , Mucosa Intestinal/irrigación sanguínea , Mucosa Intestinal/patología , Masculino , Polietilenglicoles/farmacología , Pregnatrienos/farmacología , Ratas , Ratas Sprague-Dawley , Esteroides/farmacología , Superóxido Dismutasa/farmacología
6.
J Am Vet Med Assoc ; 208(3): 404-7, 1996 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-8575974

RESUMEN

A 6-year-old 895-kg Angus bull was evaluated for a 6-month history of left hind limb lameness that was refractory to antimicrobial treatment. On physical examination, there was soft-tissue swelling associated with the lateral digit. Radiography revealed septic arthritis of the distal interphalangeal joint and osteomyelitis of the distal and middle phalanges. Treatment included debridement and lavage. Bacteriologic culture of debrided tissues yielded aerobic and anaerobic organisms. Antimicrobials were administered parenterally and locally in the form of antimicrobial-impregnated polymethyl methacrylate beads. The limb also was placed in a cast to promote ankylosis. The bull recovered, and the digit was salvaged.


Asunto(s)
Antibacterianos/administración & dosificación , Artritis Infecciosa/veterinaria , Enfermedades de los Bovinos/tratamiento farmacológico , Pezuñas y Garras , Osteomielitis/veterinaria , Actinomyces/aislamiento & purificación , Actinomicosis/tratamiento farmacológico , Actinomicosis/veterinaria , Administración Oral , Animales , Antibacterianos/uso terapéutico , Antiinflamatorios no Esteroideos/administración & dosificación , Antiinflamatorios no Esteroideos/uso terapéutico , Artritis Infecciosa/complicaciones , Artritis Infecciosa/tratamiento farmacológico , Bovinos , Cefazolina/administración & dosificación , Cefazolina/uso terapéutico , Cefalosporinas/administración & dosificación , Cefalosporinas/uso terapéutico , Clortetraciclina/administración & dosificación , Clortetraciclina/uso terapéutico , Desbridamiento/veterinaria , Enfermedades del Pie/tratamiento farmacológico , Enfermedades del Pie/veterinaria , Miembro Posterior , Masculino , Metilmetacrilatos , Metronidazol/administración & dosificación , Metronidazol/uso terapéutico , Microesferas , Osteomielitis/complicaciones , Osteomielitis/tratamiento farmacológico , Fenilbutazona/administración & dosificación , Fenilbutazona/uso terapéutico , Irrigación Terapéutica/veterinaria
7.
J Am Vet Med Assoc ; 210(7): 939-41, 1997 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-9096724

RESUMEN

Urethral obstruction is common in castrated male small ruminants. Perineal urethrostomy is the recommended treatment in animals not intended for breeding; however, stricture formation is a common complication, and few surgical options are available once a stricture has formed. Prepubic urethrostomy has been used successfully in dogs and cats to treat urethral trauma or stricture formation, and in this report, use of prepubic urethrostomy in 2 small ruminants to treat urethral strictures that developed alter perineal urethrostomy is described. Both animals maintained urinary continence and were able to void urine normally. Complications after prepubic urethrostomy included recurrent cystitis in sheep and urethral stricture in goats. Prepubic urethrostomy may be considered in as an alternative for treatment of urethral disorders in small ruminants.


Asunto(s)
Enfermedades de las Cabras/cirugía , Enfermedades de las Ovejas/cirugía , Uretra/cirugía , Obstrucción Uretral/veterinaria , Animales , Cabras , Masculino , Reoperación/veterinaria , Ovinos , Obstrucción Uretral/cirugía
8.
J Am Vet Med Assoc ; 202(2): 301-3, 1993 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-8428841

RESUMEN

Cholelithiasis is the most common cause of biliary obstruction in horses. Proposed mechanisms include ascariasis, biliary stasis, ascending biliary infection, and changes in bile composition. In this horse, a foreign body acted as the nidus for bile-salt deposition and ascending cholangitis. Clinical signs (intermittent abdominal pain, icterus, and pyrexia) in conjunction with high serum activity of enzymes indicative of obstructive biliary disease led to a tentative diagnosis of cholelithiasis. Ultrasonography was used to confirm the diagnosis. Postmortem examination revealed a 7-cm wooden stick to be the core of a cholelith found in the common bile duct.


Asunto(s)
Conducto Colédoco , Cuerpos Extraños/veterinaria , Cálculos Biliares/veterinaria , Enfermedades de los Caballos/etiología , Animales , Cuerpos Extraños/complicaciones , Cuerpos Extraños/diagnóstico por imagen , Cálculos Biliares/diagnóstico por imagen , Cálculos Biliares/etiología , Enfermedades de los Caballos/diagnóstico por imagen , Caballos , Masculino , Ultrasonografía
9.
J Am Vet Med Assoc ; 202(7): 1119-22, 1993 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-8473227

RESUMEN

A modification of the Roux-en-Y anastomosis procedure was used to bypass a pyloroduodenal mass in a 12-year-old Arabian stallion. Clinical signs had consisted of a 4-week progression of ventral and hind limb edema, hypoproteinemia, fecal occult blood, intermittent abdominal pain, weight loss, and gastric reflux. On exploratory celiotomy, an obstructive mass was found in the pylorus and proximal portion of the duodenum. Gastrojejunostomy and duodenojejunostomy were performed by use of stapled side-to-side anastomosis techniques. Inaccessibility of the obstructed pyloric region prevented resection of the affected area.


Asunto(s)
Obstrucción Duodenal/veterinaria , Duodeno/cirugía , Enfermedades de los Caballos/cirugía , Yeyuno/cirugía , Estómago/cirugía , Anastomosis en-Y de Roux/veterinaria , Animales , Obstrucción Duodenal/cirugía , Caballos , Masculino , Antro Pilórico/cirugía
10.
Can Vet J ; 38(9): 561-3, 1997 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-9285136

RESUMEN

A yearling horse was presented with a septic, transverse fracture in the frontal plane of the distal phalanx. The fracture fragment was surgically removed from the parent bone via the sole. The foot was managed by corrective farriery and the horse was able to be ridden for pleasure.


Asunto(s)
Traumatismos de los Pies/veterinaria , Fracturas Óseas/veterinaria , Caballos/cirugía , Animales , Femenino , Traumatismos de los Pies/cirugía , Fracturas Óseas/cirugía , Miembro Posterior/lesiones , Miembro Posterior/cirugía
13.
Can Vet J ; 31(10): 705-7, 1990 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-17423679
15.
South Med J ; 68(9): 1139-41, 1975 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-1162428

RESUMEN

Reported is an atypical, severe case of Hallermann-Streiff syndrome combined with progeria, bilateral microphthalmus, cataracts, and normal chromosome count. A plea is made for study into the teratology of abnormal chemical, metabolic, and other forces that attack the early stages of the development of the human fetus and produce multiple anomalies.


Asunto(s)
Síndrome de Hallermann/complicaciones , Disostosis Mandibulofacial/complicaciones , Progeria/complicaciones , Anomalías Múltiples/etiología , Adulto , Preescolar , Enanismo/etiología , Entropión/etiología , Femenino , Síndrome de Hallermann/etiología , Humanos , Discapacidad Intelectual/etiología , Masculino , Erosión de los Dientes/etiología
16.
Vet Surg ; 26(3): 172-81, 1997.
Artículo en Inglés | MEDLINE | ID: mdl-9150554

RESUMEN

OBJECTIVE: The purpose of this study was to examine the effects of ischemia and reperfusion on the biochemical integrity of equine colonic mucosa to assess the relative roles of ischemic- and reperfusion-induced damage. STUDY DESIGN: Two hours of no-flow ischemia experimentally induced by 720 degrees counterclockwise ascending colon volvulus followed by 2 hours reperfusion after derotation. ANIMALS: Ten ponies. METHODS: Ascending colon biopsies were obtained every hour for measurement of mucosal adenosine triphosphate (ATP), water, sodium, and potassium content. Additional samples were homogenized for assay of mitochondrial respiratory function. RESULTS: ATP content diminished 92% after ischemia and recovered to only 44% of control levels (P < .001 versus controls) after 2 hours reperfusion. Reperfusion increased mucosal water and decreased sodium and potassium content for the duration of the experiment. Both NADH-(pyruvate) and FADH-linked (succinate) respiration decreased after ischemia and did not recover during reperfusion indicating electron transport chain dysfunction. CONCLUSIONS: Two hours ischemia induced severe metabolic dysfunction in equine colon mucosa which persisted throughout reperfusion. Unequivocal evidence of injury specific to reperfusion was not observed in this study suggesting that much of the damage observed during reperfusion may be a continuation of injury induced during the ischemic period and not specific to reperfusion per se. CLINICAL RELEVANCE: This study suggests that greater efforts to metabolically support ischemically injured mucosa may be an important aspect of obtaining improved survival of horses affected by ascending colon volvulus (ACV).


Asunto(s)
Adenosina Trifosfato/análisis , Colon/química , Enfermedades de los Caballos/metabolismo , Enfermedades de los Caballos/fisiopatología , Pulmón/fisiopatología , Daño por Reperfusión/veterinaria , Equilibrio Hidroelectrolítico , Adenosina Trifosfato/metabolismo , Animales , Cromatografía Líquida de Alta Presión/métodos , Cromatografía Líquida de Alta Presión/veterinaria , Colon/metabolismo , Colon/fisiopatología , Caballos , Mucosa Intestinal/química , Mucosa Intestinal/metabolismo , Obstrucción Intestinal/metabolismo , Obstrucción Intestinal/fisiopatología , Obstrucción Intestinal/veterinaria , Consumo de Oxígeno/fisiología , Potasio/análisis , Potasio/metabolismo , Daño por Reperfusión/metabolismo , Daño por Reperfusión/fisiopatología , Respiración/fisiología , Sodio/análisis , Sodio/metabolismo , Factores de Tiempo
17.
Vet Surg ; 23(6): 435-41, 1994.
Artículo en Inglés | MEDLINE | ID: mdl-7871706

RESUMEN

The purposes of this study were to mechanically determine the optimal tissue bite size and to evaluate seven suture materials at their largest commercially available size for breaking strength and stiffness using cadaveric adult equine linea alba. Soft tissues were removed from the abdominal fascia of 16 adult horses. Individual test sections were created from the entire linea alba and labeled (1 through 6) starting at the umbilicus and extending craniad. A single biomechanical test was performed on each test section. Tissue bite size (3, 6, 9, 12, 15, 18, and 21 mm) significantly altered breaking strength directly in a logarithmic fashion (P < .0001; R2 = 0.94). Tissue bite size accounted for 44% and linea alba thickness for 24% of the variability in breaking strength of the equine linea alba. The optimal tissue bite size for adult horse was 15 mm from the edge of the linea alba based on lack of significant gain in breaking strength. There were no differences in breaking strength among horses, horses weight, or left and right test sections. Test sections taken from near or at the umbilicus had greater breaking strength (P < .005) and thicker linea alba (P < .001) when compared with more cranial test sections. Linea alba thickness alone accounted for 34% of the variability in breaking strength associated with test section position. There were no differences in linea alba stiffness among tissue bite sizes. All suture loops failed before complete fascial disruption, and 52 of 56 (93%) suture loops failed at the knot.(ABSTRACT TRUNCATED AT 250 WORDS)


Asunto(s)
Músculos Abdominales/fisiología , Caballos/fisiología , Suturas/veterinaria , Soporte de Peso/fisiología , Músculos Abdominales/cirugía , Animales , Fenómenos Biomecánicos , Fascia/fisiología , Dehiscencia de la Herida Operatoria/prevención & control , Dehiscencia de la Herida Operatoria/veterinaria , Suturas/clasificación , Suturas/normas
18.
J Biol Chem ; 264(12): 6707-10, 1989 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-2468660

RESUMEN

The beta-adrenergic receptor kinase, which specifically phosphorylates the agonist-occupied beta-adrenergic receptor, is strongly inhibited by polyanions. Heparin and dextran sulfate inhibit the enzyme with an IC50 of approximately 0.15 microM. De-N-sulfated heparin is approximately 8-fold less potent. Other acid mucopolysaccharides such as heparan sulfate and chondroitin sulfates B and C are also less effective. Polyaspartic and polyglutamic acid also inhibit with IC50 values of 1.3-2 microM. Inositol hexasulfate, with an IC50 of 13 microM is approximately 270-fold more potent than inositol hexaphosphate implicating the sulfate group as a major determinant of the inhibition. The inhibition by heparin is competitive with substrate and of mixed type with respect to ATP. Polycations also inhibit receptor phosphorylation by beta-adrenergic receptor kinase. Polylysine is more effective with an IC50 of 69 microM, while spermine (990 microM) and spermidine (2570 microM) are less potent. Polylysine, spermine, and spermidine are also able to block effectively the inhibition by heparin. The identification of compounds which specifically inhibit beta-adrenergic receptor kinase should prove useful in further defining the biological role of this enzyme.


Asunto(s)
Proteínas Quinasas Dependientes de AMP Cíclico , Poliaminas , Polímeros/farmacología , Inhibidores de Proteínas Quinasas , Receptores Adrenérgicos beta/metabolismo , Animales , Bovinos , Sulfato de Dextran , Dextranos/farmacología , Heparina/farmacología , Técnicas In Vitro , Cinética , Polielectrolitos , Rodopsina/metabolismo , Segmento Externo de la Célula en Bastón/enzimología , Quinasas de Receptores Adrenérgicos beta
19.
Vet Surg ; 20(4): 240-4, 1991.
Artículo en Inglés | MEDLINE | ID: mdl-1949560

RESUMEN

Salter-Harris type I fractures of the femoral capital physis were repaired in five Holstein bulls with three 7.0 mm cannulated screws placed in lag fashion. Radiographically at months 7 to 10, the fractures were healed and there was periarticular bone production on the femoral necks and the dorsal acetabular rims. Four bulls had normal gaits, and one bull had muscle atrophy and barely detectable lameness.


Asunto(s)
Tornillos Óseos/veterinaria , Bovinos/lesiones , Cabeza Femoral/lesiones , Fracturas de Cadera/veterinaria , Animales , Bovinos/cirugía , Fracturas de Cadera/cirugía , Masculino , Resultado del Tratamiento , Cicatrización de Heridas
20.
J Biol Chem ; 268(11): 8256-60, 1993 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-8463335

RESUMEN

The beta gamma subunits of heterotrimeric G proteins play important roles in regulating receptor-stimulated signal transduction processes. Recently appreciated among these is their role in the signaling events that lead to the phosphorylation and subsequent desensitization of muscarinic cholinergic (Haga, K., and Haga, T. (1992) J. Biol. Chem. 267, 2222-2227) and beta-adrenergic (Pitcher, J. A., Inglese, J., Higgins, J. B., Arriza, J. L., Casey, P. J., Kim, C., Benovic, J. L., Kwatra, M. M., Caron, M. G., and Lefkowitz, R. J. (1992) Science 257, 1264-1267) receptors. Beta gamma mediates the membrane targeting of the beta-adrenergic receptor kinase (beta ARK), in response to receptor activation, through a specific beta ARK-beta gamma interaction. This process utilizes the membrane-anchoring properties of the isoprenylated gamma subunit of beta gamma. In the present study, we have employed three distinct approaches to identify the region within the carboxyl terminus of beta ARK which binds beta gamma and thereby results in membrane translocation. We studied the ability of beta gamma to enhance the enzymatic activity of a series of truncated mutants of bovine beta ARK1, the ability of glutathione S-transferase fusion proteins containing various lengths of the carboxyl terminus of beta ARK to bind beta gamma subunits, and the ability of synthetic peptides comprised of beta ARK sequences to inhibit beta gamma activation of beta ARK1. We find that the minimal beta gamma binding domain of beta ARK is localized to a 125-amino acid residue stretch, the distal end of which is located 19 residues from the carboxyl terminus. A single 28-mer peptide (Trp643 to Ser670) derived from this sequence effectively inhibited beta gamma activation of beta ARK1, with an IC50 of 76 microM. The identification of this "beta gamma binding domain" on beta ARK and the development of peptide inhibitors provide important tools for the study of G protein-coupled receptor desensitization, as well as for the investigation of beta gamma activation of other G protein-effector systems.


Asunto(s)
Proteínas Quinasas Dependientes de AMP Cíclico , Proteínas de Unión al GTP/metabolismo , Proteínas Quinasas/metabolismo , Animales , Sitios de Unión , Bovinos , Línea Celular , Clonación Molecular , Activación Enzimática , Escherichia coli/genética , Glutatión Transferasa/genética , Glutatión Transferasa/metabolismo , Cinética , Sustancias Macromoleculares , Metionina/metabolismo , Mutagénesis , Fosforilación , Proteínas Quinasas/genética , Proteínas Recombinantes de Fusión/metabolismo , Eliminación de Secuencia , Transducción de Señal , Radioisótopos de Azufre , Transfección , Quinasas de Receptores Adrenérgicos beta
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