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1.
Can Fam Physician ; 70(1): 33-37, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38262748
2.
Transpl Infect Dis ; 14(5): E102-6, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22931101

RESUMO

A 31-year-old man underwent living-related kidney transplantation in 2004 as a consequence of primary focal segmental glomerulosclerosis (FSGS). Four years after the transplantation, we confirmed nephrotic syndrome caused by recurrent FSGS. We performed plasmapheresis and low-density lipoprotein adsorption. We also combined steroid therapy with a reduction in the dose of tacrolimus and an increased dose of mycophenolate mofetil. The nephrotic syndrome improved dramatically with this combined therapeutic approach. However, 10 months after these treatments, he revisited our hospital because of altered consciousness. We detected multiple tumor masses in his brain that were ring enhanced on contrast magnetic resonance imaging. Consequently, we suspected primary central nervous system post-transplantation lymphoproliferative disorder (CNS-PTLD). We performed a craniotomy to biopsy the brain tumors. The biopsy specimen showed Epstein-Barr virus-associated diffuse large B-cell lymphoma. There is no definitive treatment for CNS-PTLD. Therefore, we treated the primary CNS-PTLD successfully with whole-brain radiation and discontinuation of immunosuppression therapy.


Assuntos
Doenças do Sistema Nervoso Central/radioterapia , Transplante de Rim/efeitos adversos , Transtornos Linfoproliferativos/radioterapia , Adulto , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Doenças do Sistema Nervoso Central/etiologia , Doenças do Sistema Nervoso Central/patologia , Humanos , Imunossupressores/uso terapêutico , Transtornos Linfoproliferativos/etiologia , Transtornos Linfoproliferativos/patologia , Masculino , Radiografia , Resultado do Tratamento
3.
Prostate Cancer Prostatic Dis ; 11(1): 32-9, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-17984999

RESUMO

Current existing therapies for prostate cancer eradicate the majority of cells within a tumor. However, most patients with advanced cancer still progress to androgen-independent metastatic disease that remains essentially incurable by current treatment strategies. Recent evidence has shown that cancer stem cells (CSCs) are a subset of the tumor cells that are responsible for initiating and maintaining the disease. Understanding normal stem cells and CSCs may provide insight into the origin of and new therapeutics for prostate cancer. Normal stem cells and CSCs have been identified in prostate tissue by the use of several markers or techniques. Although research on stem cells has been limited by the lack of suitable in vitro systems, recent studies show that not only primary cells but also several established cell lines may exhibit stem cell properties. This review discusses various in vitro culture systems to propagate normal prostate stem cells and prostate CSCs together with molecular markers. These in vitro cell culture models should be useful for elucidating the differentiation of prostatic epithelium and the biological features of prostate cancer.


Assuntos
Modelos Biológicos , Células-Tronco Neoplásicas/patologia , Próstata/citologia , Neoplasias da Próstata/patologia , Técnicas de Cultura de Células , Humanos , Masculino , Neoplasias da Próstata/metabolismo
4.
Leukemia ; 20(3): 485-90, 2006 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16424864

RESUMO

Among 11 JMML children, two had an abnormal karyotype, and nine had a normal karyotype at onset. In one patient with trisomy 8 and four patients with a normal karyotype, a new clone with an aberrant karyotype emerged 1-14 months after 6-mercaptopurine (6-MP) therapy as shown by G-banding analyses. Fluorescence in situ hybridization disclosed that an abnormal clone existed in approximately 3-6% of bone marrow cells at onset or before 6-MP therapy in all the four cases examined, and increased to approximately 12-90% during the treatment. In culture with granulocyte-macrophage colony-stimulating factor, cytogenetically abnormal clones that proliferated during 6-MP therapy possessed significantly less sensitivity to the antimetabolite, compared with cells that decreased in numbers after the therapy. A PTPN11 mutation was detected in all of granulocyte-macrophage colonies irrespective of karyotypic aberration in one patient, whereas approximately 80% of erythroid colonies and 20% of mixed colonies possessed neither a PTPN11 mutation nor chromosomal abnormalities. The appearance of chromosomal aberrations shown by G-banding during 6-MP therapy in some JMML cases may result, in part, from the growth of a 6-MP-refractory clone that already exists at onset. It is possible that treatment with 6-MP promotes progression of the disease.


Assuntos
Antineoplásicos/uso terapêutico , Aberrações Cromossômicas , Leucemia Mielomonocítica Aguda/tratamento farmacológico , Leucemia Mielomonocítica Aguda/genética , Mercaptopurina/uso terapêutico , Bandeamento Cromossômico , Genes ras , Humanos , Hibridização in Situ Fluorescente , Peptídeos e Proteínas de Sinalização Intracelular/genética , Leucemia Mielomonocítica Aguda/patologia , Mutação , Proteína Tirosina Fosfatase não Receptora Tipo 11 , Proteínas Tirosina Fosfatases/genética
5.
Interv Neuroradiol ; 12(Suppl 1): 86-90, 2006 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-20569608

RESUMO

SUMMARY: In this paper, we report five cases with acutely ruptured wide-necked aneurysms, which were treated with coil embolization using a balloon or stent-assisted technique. Balloon-assisted coil embolization using Equinox balloon, Commodore balloon, and Hyperform balloon were performed for four patients and stent-assisted coil embolization using BX velocity for one patient. We discuss problems of coil embolization for acutely ruptured wide-necked aneurysms with a balloon or stent-assisted technique.

6.
Curr Issues Mol Biol ; 4(4): 129-46, 2002 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-12432964

RESUMO

Human and other annotated genome sequences have facilitated generation of vast amounts of correlative data, from human/animal genetics, normal and disease-affected tissues from complex diseases such as arthritis using gene/protein chips and SNP analysis. These data sets include genes/proteins whose functions are partially known at the cellular level or may be completely unknown (e.g. ESTs). Thus, genomic research has transformed molecular biology from "data poor" to "data rich" science, allowing further division into subpopulations of subcellular fractions, which are often given an "-omic" suffix. These disciplines have to converge at a systemic level to examine the structure and dynamics of cellular and organismal function. The challenge of characterizing ESTs linked to complex diseases is like interpreting sharp images on a blurred background and therefore requires a multidimensional screen for functional genomics ("functionomics") in tissues, mice and zebra fish model, which intertwines various approaches and readouts to study development and homeostasis of a system. In summary, the post-genomic era of functionomics will facilitate to narrow the bridge between correlative data and causative data by quaint hypothesis-driven research using a system approach integrating "intercoms" of interacting and interdependent disciplines forming a unified whole as described in this review for Arthritis.


Assuntos
Biologia Computacional , Perfilação da Expressão Gênica , Genômica , Osteoartrite/genética , Animais , Cartilagem/metabolismo , Análise por Conglomerados , Colágeno/genética , Colágeno/metabolismo , Citocinas/metabolismo , Endopeptidases/metabolismo , Fibronectinas/metabolismo , Genoma , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Osteopontina , Fatores de Risco , Sialoglicoproteínas/metabolismo
7.
Hinyokika Kiyo ; 47(3): 175-7, 2001 Mar.
Artigo em Japonês | MEDLINE | ID: mdl-11329958

RESUMO

An 84-year-old man presented at our hospital with complaints of severe gross hematuria and lower right abdominal pain. A right renal mass was detected by ultrasound sonography and plain computerized tomography (CT) scan, but an exact diagnosis was not obtained. Because the patient presented with moderate renal dysfunction and severe gross hematuria, we were unable to perform imaging studies using contrast material or ureteroscopic instruments. Finally, mercaptoacetylglycyl-glycylglycine (MAG3) scintigraphy and magnetic resonance imaging (MRI) demonstrated renal cell carcinoma, and we performed transarterial embolization (TAE) therapy using ethanol and gel foam. Based on their efficacy and noninvasiveness, we conclude that MAG3 scintigraphy and MRI are the optimal modalities for imaging in patients with renal dysfunction.


Assuntos
Carcinoma de Células Renais/diagnóstico , Nefropatias/diagnóstico , Neoplasias Renais/diagnóstico , Tecnécio Tc 99m Mertiatida , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Renais/terapia , Embolização Terapêutica , Etanol/administração & dosagem , Humanos , Nefropatias/complicações , Neoplasias Renais/terapia , Imageamento por Ressonância Magnética , Masculino
8.
J Neurol Sci ; 184(2): 189-96, 2001 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-11239955

RESUMO

We report an elderly woman with multiple sclerosis who showed an extensive cavity formation in the midthoracic cord in addition to multiple abnormal intensity signals in the central nervous system on magnetic resonance imaging (MRI). The cavity decreased in size in response to corticosteroid therapy with an improvement in neurological symptoms. The autopsy demonstrated a slit-like cavity lined with no ependymal cells on the luminal surface in the lower cervical to midthoracic cord, with circumferentially distributed demyelinative lesions, leading to the pathological diagnosis of secondary syringomyelia. In this patient a limited necrosis formed in the spinal cord might have developed into a cavity formation with edematous fluid leading to subsequent episodes of neurological exacerbation.


Assuntos
Tronco Encefálico/patologia , Esclerose Múltipla/patologia , Nervo Óptico/patologia , Medula Espinal/patologia , Siringomielia/patologia , Idoso , Vértebras Cervicais , Feminino , Humanos , Esclerose Múltipla/complicações , Siringomielia/complicações , Vértebras Torácicas
9.
J Neurol Sci ; 183(1): 5-12, 2001 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-11166787

RESUMO

We performed brain CT and single-photon emission computed tomography (SPECT) using N-isopropyl-p-[123I] iodoamphetamine (123I-IMP) as a tracer in the early stage of seven patients with Creutzfeldt-Jakob disease (CJD). In four of the patients, we determined absolute values of regional cerebral blood flow (rCBF) in the frontal, temporal, parietal and occipital lobes, thalamus and cerebellum using an autoradiographic method with a single blood sample. Brain CT demonstrated no abnormal findings other than a mild age-related atrophy in all patients except for one patient with a low-density area in the left cerebellar hemisphere due to an old hemorrhage, whereas SPECT revealed a decreased uptake of the tracer in various parts of the cerebral cortex of all patients, sometimes in an asymmetrical pattern. Absolute values of rCBF showed a significant decrease in all examined regions of the patients as against healthy controls (P<0.0001). In three patients, SPECT demonstrated a decreased uptake throughout the cerebral cortex on visual inspection, whereas absolute values of rCBF revealed an obvious decrease of the uptake also in the thalamus and cerebellum. These results suggest that SPECT with quantification of rCBF using 123I-IMP might be a sensitive and useful technique not only for detecting a focal metabolic dysfunction but also for diagnosis in the early stage of CJD.


Assuntos
Encéfalo/irrigação sanguínea , Síndrome de Creutzfeldt-Jakob/diagnóstico por imagem , Iofetamina , Compostos Radiofarmacêuticos , Tomografia Computadorizada de Emissão de Fóton Único , Idoso , Idoso de 80 Anos ou mais , Encéfalo/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estatísticas não Paramétricas , Tomografia Computadorizada por Raios X
10.
Intern Med ; 40(11): 1149-53, 2001 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-11757774

RESUMO

We report a 63-year-old man with acute disseminated encephalomyelitis (ADEM), initially showing depression for one and a half months but subsequently meningoencephalitis followed by acute-onset myelopathy. Neuroradiological examinations of the brain demonstrated no focal lesion causative for his depression, while cerebrospinal fluid revealed elevated levels of inflammatory cytokines in parallel with disease activity. Because depression is usually a rare initial symptom for patients with ADEM, an increased production of inflammatory cytokines in the central nervous system as well as age-related alterations of immune response might have played an important role in the development of depression in this elderly patient.


Assuntos
Depressão/etiologia , Encefalomielite Aguda Disseminada/complicações , Encefalomielite Aguda Disseminada/diagnóstico , Líquido Cefalorraquidiano/metabolismo , Depressão/líquido cefalorraquidiano , Encefalomielite Aguda Disseminada/líquido cefalorraquidiano , Humanos , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença
11.
J Biochem ; 128(4): 629-35, 2000 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-11011145

RESUMO

Two monoclonal antibodies, beta 208 and beta 210, against the beta subunit of the F(1) ATPase from Escherichia coli reacted with an intact beta subunit and also a peptide corresponding to a portion of beta between residues 1 and 145. Mutations at Ala-1, Val-15, Glu-16, Phe-17, Leu-29, Gly-65, or Leu-66, and His-110 or Arg-111 for beta 210 and beta 208, respectively, caused decreased antibody binding to beta, suggesting that these residues form the epitopes and are thought to lie close together on the surface of the beta subunit. The topological locations of the corresponding residues in the atomic structure of the bovine beta subunit agree well with these expectations, except for Ala-1 and Leu-29. beta 210 binds to two beta strands including the epitope residues that are 50 residues apart, indicating that this antibody recognizes the tertiary structure of the N-terminal end region. Mutations in the epitope residues of beta 210 do not affect the F(1) ATPase activity, suggesting that surfaces of the two beta strands in the amino-terminal end region are not functionally essential. To analyze the functional importance around His-110 recognized by beta 208 we introduced site specific mutations at residues His-110 and Ile-109. Ile-109 to Ala or Arg, and His-110 to Ala or Asp caused defective assembly of F(1). However, the His-110 to Arg mutation had no effect on molecular assembly, suggesting that Ile-109 and His-110, especially the positive charge of His-110 are essential for the assembly of F(1). The His-110 to Arg mutation caused a large decrease in F(1)-ATPase activity, suggesting that a subtle change in the topological arrangement of the positive charge of His-110 located on the surface of beta plays an important role in the catalytic mechanism of the F(1)-ATPase.


Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Superfície/imunologia , Epitopos/imunologia , Epitopos/metabolismo , Escherichia coli/enzimologia , ATPases Translocadoras de Prótons/química , ATPases Translocadoras de Prótons/imunologia , Sequência de Aminoácidos , Substituição de Aminoácidos , Animais , Reações Antígeno-Anticorpo , Antígenos de Superfície/química , Antígenos de Superfície/genética , Antígenos de Superfície/metabolismo , Catálise , Sequência Conservada , Mapeamento de Epitopos , Epitopos/química , Epitopos/genética , Camundongos , Camundongos Endogâmicos BALB C , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Mutação , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína , Subunidades Proteicas , ATPases Translocadoras de Prótons/genética , ATPases Translocadoras de Prótons/metabolismo
12.
Nihon Naika Gakkai Zasshi ; 89(7): 1392-7, 2000 Jul 10.
Artigo em Japonês | MEDLINE | ID: mdl-10934768
13.
Rinsho Shinkeigaku ; 40(2): 160-5, 2000 Feb.
Artigo em Japonês | MEDLINE | ID: mdl-10835938

RESUMO

We report a case of motor neuron disease (MND) with dementia, presenting motor aphasia as an initial symptom. A 67-year-old man was admitted to our hospital because of speech disturbance slowly progressing for 2 years. On physical examination, he showed no neurological abnormalities except for non-fluent aphasia and increased deep tendon reflexes without laterality. MRI demonstrated bilateral fronto-temporal atrophy, dominating the left hemisphere. This finding was confirmed by surface anatomy scanning (SAS), showing an obvious atrophy in the left inferior frontal gyrus, compared with the right one. SPECT with 123I-IMP revealed some irregular defects in the bilateral frontotemporal region. Because he showed dementia, bulbar palsy with tongue atrophy, weakness of upper extremities and facial muscles, snout reflex, and the atrophy and fasciculation in limbs in addition to motor aphasia soon after the discharge from our hospital, he was diagnosed as having MND with dementia. At age 68, he died of a respiratory failure 3 years after the onset of the disease. MND with dementia seldom shows motor aphasia as an initial symptom. We must include, however, the MND with dementia as an differential diagnosis when we see the patients with progressive aphasia.


Assuntos
Afasia de Broca/etiologia , Demência/complicações , Doença dos Neurônios Motores/complicações , Idoso , Atrofia , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Paralisia Bulbar Progressiva/complicações , Diagnóstico Diferencial , Humanos , Imageamento por Ressonância Magnética , Masculino , Doença dos Neurônios Motores/diagnóstico , Tomografia Computadorizada de Emissão de Fóton Único
14.
Intern Med ; 39(6): 490-4, 2000 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-10852171

RESUMO

A 46-year-old woman showed proteinuria and hematuria after left blepharoptosis, and revealed a histopathology of membranous nephropathy (MN) at renal biopsy. She was diagnosed as having myasthenia gravis (MG) because of a positive edrophonium test and anti-acetylcholine receptor (AchR) antibodies in serum. We found a decrease in anti-AchR antibodies after extended total thymectomy, in parallel with an improvement in both urinary findings and myasthenic symptoms. In this case, MG preceded MN and the thymectomy was effective for both diseases, suggesting that the thymus might play an important role in the pathogenesis of MN.


Assuntos
Glomerulonefrite Membranosa/complicações , Glomerulonefrite Membranosa/cirurgia , Miastenia Gravis/complicações , Miastenia Gravis/cirurgia , Timectomia/métodos , Feminino , Humanos , Pessoa de Meia-Idade , Indução de Remissão
15.
Rinsho Ketsueki ; 40(7): 568-73, 1999 Jul.
Artigo em Japonês | MEDLINE | ID: mdl-10483140

RESUMO

An 11-month-old boy was transferred to our hospital because of fever and bleeding tendency on March 13, 1998. Laboratory studies showed a white blood cell count of 43,360/microliter with 75% blasts, a hemoglobin concentration of 8.4 g/dl, and a platelet count of 23 x 10(3)/microliter. Surface marker analysis with a flow cytometer revealed that only 21% and 11% of the blasts, respectively, were positive for CD41 and CD42b. Treatment with a permeabilizing agent apparently increased the reactivity of the blasts with anti-CD41 monoclonal antibody (MoAb), which can recognize IIb independently of IIIa. However no significant differences were observed in reactivity with anti-CD41 MoAb (which recognizes the IIb/IIIa complex) anti-CD61 MoAb and anti-CD42b MoAb before or after fixation. Blasts positive for platelet peroxidase were observed by electron microscopy, thus confirming the diagnosis of acute megakaryoblastic leukemia. We concluded that the detection of intracellular antigens is useful for the quick diagnosis of acute megakaryoblastic leukemia characterized by low surface expression of megakaryocytic lineage antigens.


Assuntos
Leucemia Megacarioblástica Aguda/diagnóstico , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/análise , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Citarabina/administração & dosagem , Etoposídeo/administração & dosagem , Humanos , Lactente , Leucemia Megacarioblástica Aguda/tratamento farmacológico , Masculino , Megacariócitos/imunologia , Mitoxantrona/administração & dosagem , Indução de Remissão
16.
J Clin Endocrinol Metab ; 84(1): 363-6, 1999 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-9920108

RESUMO

Autosomal dominant hypocalcemia (ADH), caused by activating mutations of the calcium-sensing receptor (CaSR), is characterized by hypocalcemia with an inappropriately low concentration of PTH. Among 11 missense mutations of CaSR reported to date in patients with ADH or sporadic hypocalcemia, functional properties of 8 mutant CaSRs were characterized. Here, we describe a novel mutation of CaSR and its functional property in a family with ADH. The 41-yr-old male proband had asymptomatic hypocalcemia with a history of recurrent nephrolithiasis. His father had asymptomatic hypocalcemia, but his mother was normocalcemic. PCR-single strand conformation polymorphism and sequencing revealed that both the proband and the father had a novel heterozygous mutation in CaSR gene that causes lysine to asparagine substitution at codon 47 (K47N), which is in the extracellular domain of CaSR, like 6 of 11 known activating mutations. Using HEK293 cells transfected with wild-type or K47N CaSR complementary DNA, the intracellular Ca2+ concentration was assessed in response to changes in the extracellular Ca2+ concentration. The EC50 of the mutant CaSR for the extracellular Ca2+ concentration was 2.2 mmol/L and was significantly lower than that of wild-type (3.7 mmol/L). These results confirm that this mutation is responsible for ADH in this family. The fact that several inactivating mutations in familial hypocalciuric hypercalcemia occur in amino acid around K47 suggests the importance of the N-terminal portion of the receptor in extracellular Ca sensing.


Assuntos
Hipocalcemia/genética , Mutação de Sentido Incorreto , Receptores de Superfície Celular/genética , Adulto , DNA/química , Genes Dominantes , Humanos , Masculino , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , Receptores de Detecção de Cálcio
17.
Mov Disord ; 13(6): 934-40, 1998 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-9827618

RESUMO

We studied the clinical efficacy of mexiletine, a derivative oral form of lidocaine, for treatment of spasmodic torticollis. One of the nine subjects of this study was previously reported. Before starting oral mexiletine, normal saline was first injected intravenously as placebo control; lidocaine infusion then followed and clinical evaluation was provided by dystonia rating scale scores, videotape recordings, and surface electromyographic recording. In all patients, lidocaine injection resulted in a decrease of dystonic muscle contractions within 5 minutes and the effect lasted approximately 1 hour. With gradual increase of mexiletine dose, similar clinical improvement was obtained with oral doses ranging from 450-1200 mg/day for more than 6 months. Side effects in six of nine patients included upper gastrointestinal symptoms, dizziness, ataxia, and dysarthria. These were tolerable or medically manageable; only one patient required a small reduction in mexiletine dose. Strong positive correlation was found between serum and cerebrospinal fluid (CSF) mexiletine concentrations with a CSF/serum ratio of 0.6 (r = 0.96, p = 0.0005) suggesting its effective penetrance into the central nervous system. We suggest that oral mexiletine therapy may be a safe and effective treatment for spasmodic torticollis.


Assuntos
Anestésicos Locais/uso terapêutico , Mexiletina/uso terapêutico , Torcicolo/tratamento farmacológico , Administração Oral , Adulto , Análise de Variância , Distonia/tratamento farmacológico , Eletromiografia , Feminino , Humanos , Infusões Intravenosas , Lidocaína/uso terapêutico , Masculino , Pessoa de Meia-Idade , Inquéritos e Questionários
19.
Arch Biochem Biophys ; 317(2): 348-56, 1995 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-7893149

RESUMO

Five monoclonal antibodies against the alpha subunit of F1-ATPase from Escherichia coli alpha 104, alpha 105, alpha 107, alpha 109, and alpha 110 were prepared. The monoclonal antibodies alpha 104 and alpha 110 enhanced the F1-ATPase activity maximally to 1.6- and 1.7-fold that of the wild-type, respectively, while alpha 105 did not. Both antibodies bound to a peptide corresponding to the region between residues 354 and 513. Mutations in this region which caused reduced binding of the alpha subunit to the antibodies were identified at residues Ser-440, Leu-456, Leu-471, Leu-482, Met-483, and Ser-506 for alpha 104 and residues Ser-440, Leu-456, Leu-471, Asp-476, Leu-482, Met-483, and Ser-506 for alpha 110. These residues are possibly involved in the epitopes for the antibodies and are located close together on the surface of the alpha subunit. Among the mutations, Leu-456 to Pro and Ser-440 to Pro mutations caused increase of the F1-ATPase activity up to 1.9 and 1.2 times that of the wild-type, respectively, while Leu-471 to Pro mutation caused a defect in assembly of the F1-ATPase on the membrane. The other mutations caused no significant change in ATPase activity. These results suggested that Ser-440 and Leu-456 have an important role in regulating catalysis by the F1-ATPase, but that the neighboring residue Leu-471 has an important role in assembly of the F1-ATPase complex. It was also suggested that binding of the monoclonal antibodies alpha 104 and alpha 110 to residues Ser-440 and Leu-456 caused local conformational changes, leading to enhancing effects on F1-ATPase activity similar to the Ser-440 to Pro and Leu-456 to Pro mutations.


Assuntos
Anticorpos Monoclonais/metabolismo , Escherichia coli/enzimologia , Leucina , ATPases Translocadoras de Prótons/química , Serina , Animais , Anticorpos Monoclonais/farmacologia , Sequência de Bases , Sítios de Ligação de Anticorpos , Western Blotting , Escherichia coli/genética , Substâncias Macromoleculares , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Mutagênese , Mapeamento de Peptídeos , Reação em Cadeia da Polimerase , ATPases Translocadoras de Prótons/genética , ATPases Translocadoras de Prótons/metabolismo , Relação Estrutura-Atividade
20.
Biochim Biophys Acta ; 1187(1): 67-72, 1994 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-8061038

RESUMO

The mutation of serine-174 to phenylalanine that causes a defect in the Escherichia coli F1-ATPase beta-subunit is suppressed by further mutations; Gly-149 to Ser, Ala-295 to Thr, Ala-295 to Pro, or Leu-400 to Gln (Miki, J., Fujiwara, K., Tsuda, M., Tsuchiya, T. and Kanazawa, H. (1990) J. Biol. Chem. 265, 21567-21572). We analyzed the effects of these second site mutations and of a newly identified Asn-158 to Tyr mutation on the activities of the ATPase without the original Ser-174 to Phe mutation. The beta-subunit with each amino acid replacement was expressed in the mutant strain JP17, which does not have a beta-subunit. Cells transformed with the plasmid carrying Ala-295 to Pro mutation alone did not grow on minimal medium agar supplemented with succinate as the sole carbon source, and showed 3% of the wild-type ATPase activity, suggesting that this mutation caused structural alterations affecting the catalytic function of the enzyme. Conversely transformants with other mutations grew well and had higher ATPase activities, suggesting that these mutations did not cause extensive structural alterations. From the transformants with the plasmid carrying the Ala-295 to Pro mutation, seven revertants capable of cell growth on succinate plates were isolated and reversion mutations were identified at residues 140, 159, 166, 171, 172 and 184 of the beta-subunits. The results suggested that Ser-174 and Ala-295 do not necessarily interact directly, but that the regions including these suppression mutation sites close to Ser-174, and Ala-295 interact with each other for the proper functioning of the ATPase. The ternary structure of the region surrounded by the residues which were identified as the reversion mutation sites for Ser-174 to Phe and Ala-295 to Pro mutations is important for the catalytic function of this enzyme.


Assuntos
Alanina/química , Escherichia coli/enzimologia , ATPases Translocadoras de Prótons/química , Serina/química , Sequência de Bases , Sítios de Ligação , Escherichia coli/genética , Dados de Sequência Molecular , Estrutura Molecular , Mutação Puntual , ATPases Translocadoras de Prótons/genética
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