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Tyrosine hydroxylase deficiency: a treatable disorder of brain catecholamine biosynthesis

WILLEMSEN, Michel A.; VERBEEK, Marcel M.; KAMSTEEG, Erik-Jan; ANDEL, Johanneke F. de Rijk-van; AEBY, Alec; BLAU, Nenad; BURLINA, Alberto; DONATI, Maria A.; GEURTZ, Ben; GRATTAN-SMITH, Padraic J.; HAEUSSLER, Martin; HOFFMANN, Georg F.; JUNG, Hans; KLERK, J
Fonte: OXFORD UNIV PRESS Publicador: OXFORD UNIV PRESS
Tipo: Artigo de Revista Científica
ENG
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36.59%
Tyrosine hydroxylase deficiency is an autosomal recessive disorder resulting from cerebral catecholamine deficiency. Tyrosine hydroxylase deficiency has been reported in fewer than 40 patients worldwide. To recapitulate all available evidence on clinical phenotypes and rational diagnostic and therapeutic approaches for this devastating, but treatable, neurometabolic disorder, we studied 36 patients with tyrosine hydroxylase deficiency and reviewed the literature. Based on the presenting neurological features, tyrosine hydroxylase deficiency can be divided in two phenotypes: an infantile onset, progressive, hypokinetic-rigid syndrome with dystonia (type A), and a complex encephalopathy with neonatal onset (type B). Decreased cerebrospinal fluid concentrations of homovanillic acid and 3-methoxy-4-hydroxyphenylethylene glycol, with normal 5-hydroxyindoleacetic acid cerebrospinal fluid concentrations, are the biochemical hallmark of tyrosine hydroxylase deficiency. The homovanillic acid concentrations and homovanillic acid/5-hydroxyindoleacetic acid ratio in cerebrospinal fluid correlate with the severity of the phenotype. Tyrosine hydroxylase deficiency is almost exclusively caused by missense mutations in the TH gene and its promoter region...

Implementação da análise de acoplamentos estatísticos e sua aplicação à família de proteínas tirosina fosfatases; Implementation of the statistical coupling analysis and its application to the Protein Tyrosine Phosphatases family.

Bleicher, Lucas
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Tese de Doutorado Formato: application/pdf
Publicado em 09/03/2009 PT
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36.57%
A Análise de Acoplamentos Estatísticos é uma técnica computacional capaz de identificar resíduos importantes para a estrutura e função de proteínas em uma família por meio da quantificação de conservação posicional, correlação entre posições e identificação de grupos de resíduos correlacionados entre si. Neste trabalho, a análise de acoplamentos estatísticos foi implementada e aplicada ao estudo das proteínas tirosina fosfatases. Em conjunto com as proteínas tirosina quinases (PTKs), que adicionam um grupo fosforil a um resíduo de tirosina em uma proteína, as proteínas tirosina fosfatases (PTPs), que o removem, são responsáveis por diversos processos de sinalização celular. Elas são um caso de evolução convergente, onde um subgrupo (as proteínas tirosina fosfatases de baixo peso molecular) não apresenta homologia às chamadas PTPs "clássicas", capazes de defosforilar apenas resíduos de tirosina, e às fosfatases de especifidicade dupla, capazes de defosforilar também resíduos de serina e treonina, além de substratos não-protéicos. Em comum, as três subfamílias apresentam apenas o motivo CX5R, característico para todas as PTPs. Através do estudo das três subfamílias utilizando a análise de acoplamentos estatísticos...

Efeitos dos inibidores de tirosina-quinase sobre a maquinaria apoptótica na leucemia mielóide crônica; The effect of tyrosine-kinase inhibitors on the apoptosis machinery in chronic myeloid leukemia

Ferreira, Aline Fernanda
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 20/12/2007 PT
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36.54%
A leucemia mielóide crônica (LMC) é uma doença mieloproliferativa, resultante da expansão clonal da célula-tronco hematopoética pluripotente. A fisiopatologia da LMC está associada a uma translocação entre os braços longos dos cromossomos 9 e 22, o que promove o aparecimento do neogene bcr-abl, cujo gene codifica uma proteína denominada Bcr-Abl. A oncoproteína Bcr-Abl possui atividade tirosina-quinase constitutiva que é a responsável pelo fenótipo maligno da célula, incluindo resistência à apoptose. O tratamento da LMC pode ser realizado com hidroxiuréia, IFN- associado à citarabina, inibidores de TK (mesilato de imatinibe e dasatinibe) e transplante de medula óssea. O tratamento de escolha para pacientes com LMC na fase crônica é o inibidor de tirosina-quinase mesilato de imatinibe e para os refratários utiliza-se o dasatinibe. Apesar do conhecimento acerca do mecanismo de ação dos inibidores de TK, pouco se sabe sobre seu efeito na maquinaria apoptótica. Sendo assim, no presente trabalho foi detectada a expressão dos genes e proteínas anti- (A1, Bcl-2, Bcl-Xl, Bcl-W, C-Flip, Ciap-1, Ciap-2 e Mcl-1) e pró-apoptóticos (Bad, Bak, Bax, Bcl-Xs, Bid, Bik, Bimel, Bmf, Bok, Fas, Fasl, Noxa e Puma) em células mononucleares de 32 indivíduos saudáveis e 26 pacientes com LMC antes e após 12 meses da terapia com mesilato de imatinibe e dasatinibe. Dentre os 26 pacientes avaliados...

Heterologous production of caffeic acid from tyrosine in Escherichia coli

Rodrigues, Joana Lúcia; Araújo, R. G.; Prather, Kristala L. J.; Kluskens, Leon; Rodrigues, L. R.
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
Publicado em //2015 ENG
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36.52%
Caffeic acid is a plant secondary metabolite and its biological synthesis has attracted increased attention due to its beneficial effects on human health. In this study, Escherichia coli was engineered for the production of caffeic acid using tyrosine as the initial precursor of the pathway. The pathway design included tyrosine ammonia lyase (TAL) from Rhodotorula glutinis to convert tyrosine to p-coumaric acid and 4-coumarate 3-hydroxylase (C3H) from Saccharothrix espanaensis or cytochrome P450 CYP199A2 from Rhodopseudomonas palustris to convert p-coumaric acid to caffeic acid. The genes were codon-optimized and different combinations of plasmids were used to improve the titer of caffeic acid. TAL was able to efficiently convert 3 mM of tyrosine to p-coumaric acid with the highest production obtained being 2.62 mM (472 mg/L). CYP199A2 exhibited higher catalytic activity towards p-coumaric acid than C3H. The highest caffeic acid production obtained using TAL and CYP199A2 and TAL and C3H was 1.56 mM (280 mg/L) and 1 mM (180 mg/L), respectively. This is the first study that shows caffeic acid production using CYP199A2 and tyrosine as the initial precursor. This study suggests the possibility of further producing more complex plant secondary metabolites like flavonoids and curcuminoids.

Tyrosine promotes oxidative stress in cerebral cortex of young rats

Sgaravatti, ??ngela Malysz; Vargas, B??thania Andrade de; Zandona, Bernardo Remuzzi; Deckamann, K??tia Bueno; Rockenbach, Francieli Juliana; Moraes, Tarsila Barros; Monserrat, Jos?? Mar??a; Sgarbi, Miriam; Pederzolli, Carolina Didonet; Wyse, Angela Terezi
Fonte: Universidade Federal do Rio Grande Publicador: Universidade Federal do Rio Grande
Tipo: Artigo de Revista Científica
ENG
Relevância na Pesquisa
36.57%
Tyrosine accumulates in inborn errors of tyrosine catabolism, especially in tyrosinemia type II, where tyrosine levels are highly elevated in tissues and physiological fluids of affected patients. In tyrosinemia type II, high levels of tyrosine are correlated with eyes, skin and central nervous system disturbances. Considering that the mechanisms of brain damage in these disorders are poorly known, in the present study, we investigated whether oxidative stress is elicited by L-tyrosine in cerebral cortex homogenates of 14-day-old Wistar rats. The in vitro effect of 0.1???4.0 mM Ltyrosine was studied on the following oxidative stress parameters: total radical-trapping antioxidant potential (TRAP), total antioxidant reactivity (TAR), ascorbic acid content, reduced glutathione (GSH) content, spontaneous chemiluminescence, thiobarbituric acid-reactive substances (TBARS), thiol-disulfide redox state (SH/SS ratio), protein carbonyl content, formation of DNA-protein cross-links, and the activities of the enzymes superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glucose-6-phosphate dehydrogenase (G6PDH). TRAP, TAR,ascorbic acid content, SH/SS ratio and CAT activity were significantly diminished, while formation of DNA-protein cross-link was significantly enhanced by L-tyrosine in vitro. In contrast...

Mutational analysis of the carboxy-terminal (YGX)4 repeat domain of CpsD, an autophosphorylating tyrosine kinase required for capsule biosynthesis in Streptococcus pneumoniae

Morona, J.; Morona, R.; Miller, D.; Paton, J.
Fonte: Amer Soc Microbiology Publicador: Amer Soc Microbiology
Tipo: Artigo de Revista Científica
Publicado em //2003 EN
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36.64%
In Streptococcus pneumoniae, CpsB, CpsC, and CpsD are essential for encapsulation, and mutants containing deletions of cpsB, cpsC, or cpsD exhibit rough colony morphologies. CpsD is an autophosphorylating protein-tyrosine kinase, CpsC is required for CpsD tyrosine phosphorylation, and CpsB is a phosphotyrosine-protein phosphatase. We have previously shown that autophosphorylation of CpsD at tyrosine attenuates its activity and consequently reduces the level of encapsulation and negatively regulates CPS production. In this study, we further investigated the role of the carboxy-terminal (YGX)4 repeat domain of CpsD in encapsulation. A CpsD truncation mutant in which the entire (YGX)4 repeat domain was removed was indistinguishable from a strain in which the entire cpsD gene had been deleted, indicating that the carboxy-terminal (YGX)4 tail is required for CpsD activity in capsular polysaccharide production. Double mutants having a single tyrosine residue at position 2, 3, or 4 in the (YGX)4 repeat domain and lacking CpsB exhibited a rough colony morphology, indicating that in the absence of an active protein-tyrosine phosphatase, phosphorylation of just one of the tyrosine residues in the (YGX)4 repeat was sufficient to inactivate CpsD. When various mutants in which CpsD had either one or combinations of two or three tyrosine residues in the (YGX)4 repeat domain were examined...

Src homology 2 domain-containing protein-tyrosine phosphatases, SHP-1 and SHP-2, are required for platelet endothelial cell adhesion molecule-1/CD31-mediated inhibitory signaling

Henshall, T.; Jones, K.; Wilkinson, R.; Jackson, D.
Fonte: Amer Assoc Immunologists Publicador: Amer Assoc Immunologists
Tipo: Artigo de Revista Científica
Publicado em //2001 EN
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36.63%
Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) is a newly assigned member of the Ig immunoreceptor tyrosine-based inhibitory motif superfamily, and its functional role is suggested to be an inhibitory receptor that modulates immunoreceptor tyrosine-based activation motif-dependent signaling cascades. To test whether PECAM-1 is capable of delivering inhibitory signals in B cells and the functional requirement of protein-tyrosine phosphatases (PTPs) for this inhibitory signaling, we generated chimeric Fc gamma RIIB1-PECAM-1 receptors containing the extracellular and transmembrane portions of murine Fc gamma RIIB1 and the cytoplasmic domain of human PECAM-1. These chimeric receptors were stably expressed in chicken DT40 B cells either as wild-type or mutant cells deficient in SHP-1(-/-), SHP-2(-/-), SHIP(-/-), or SHP-1/2(-/-) and then assessed for their ability to inhibit B cell Ag receptor (BCR) signaling. Coligation of wild-type Fc gamma RIIB1-PECAM-1 with BCR resulted in inhibition of intracellular calcium release, suggesting that the cytoplasmic domain of PECAM-1 is capable of delivering an inhibitory signal that blocks BCR-mediated activation. This PECAM-1-mediated inhibitory signaling correlated with tyrosine phosphorylation of the Fc gamma RIIB1-PECAM-1 chimera...

Tyrosine phosphorylation of CpsD negatively regulates capsular polysaccharide biosynthesis in Streptococcus pneumoniae

Morona, J.; Paton, J.; Miller, D.; Morona, R.
Fonte: Blackwell Publishing Ltd Publicador: Blackwell Publishing Ltd
Tipo: Artigo de Revista Científica
Publicado em //2000 EN
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36.57%
In Streptococcus pneumoniae, the first four genes of the capsule locus (cpsA to cpsD) are common to most serotypes. By analysis of various in-frame deletion and site-directed mutants, the function of their gene products in capsular polysaccharide (CPS) biosynthesis was investigated. We found that while CpsB, C and D are essential for encapsulation, CpsA is not. CpsC and CpsD have similarity to the amino-terminal and carboxy-terminal regions, respectively, of the autophosphorylating protein-tyrosine kinase Wzc from Escherichia coli. Alignment of CpsD with Wzc and other related proteins identified conserved Walker A and B sequence motifs and a tyrosine rich domain close to the carboxy-terminus. We have shown that CpsD is also an autophosphorylating protein-tyrosine kinase and that point mutations in cpsD affecting either the ATP-binding domain (Walker A motif) or the carboxy-terminal [YGX]4 repeat domain eliminated tyrosine phosphorylation of CpsD. We describe, for the first time, the phenotypic impact of these two mutations on polysaccharide production and show that they affect CPS production differently. Whereas a mutation in the Walker A motif resulted in loss of encapsulation, mutation of the tyrosines in the [YGX]4 repeat domain resulted in an apparent increase in encapsulation and a mucoid phenotype. These data suggest that autophosphorylation of CpsD at tyrosine attenuates its activity and reduces the level of encapsulation. Additionally...

Analysis of the mechanism by which calcium negatively regulates the tyrosine phosphorylation cascade associated with sperm capacitation

Baker, Mark A.; Hetherington, Louise; Ecroyd, Heath William; Roman, Shaun D.; Aitken, R. John
Fonte: Company of Biologists Limited Publicador: Company of Biologists Limited
Tipo: Artigo de Revista Científica
Publicado em //2004 EN
Relevância na Pesquisa
36.59%
The capacitation of mammalian spermatozoa involves the activation of a cAMP-mediated signal transduction pathway that drives tyrosine phosphorylation via mechanisms that are unique to this cell type. Controversy surrounds the impact of extracellular calcium on this process, with positive and negative effects being recorded in independent publications. We clearly demonstrate that the presence of calcium in the external medium decreases tyrosine phosphorylation in both human and mouse spermatozoa. Under these conditions, a rise in intracellular pH was recorded, however, this event was not responsible for the observed changes in phosphotyrosine expression. Rather, the impact of calcium on tyrosine phosphorylation in these cells was associated with an unexpected change in the intracellular availability of ATP. Thus, the ATP content of both human and mouse spermatozoa fell significantly when these cells were incubated in the presence of external calcium. Furthermore, the removal of glucose, or addition of 2-deoxyglucose, decreased ATP levels within human spermatozoon populations and induced a corresponding decline in phosphotyrosine expression. In contrast, the mitochondrial inhibitor rotenone had no effect on either ATP levels or tyrosine phosphorylation. Addition of the affinity-labeling probe 8-N3 ATP confirmed our prediction that spermatozoa have many calcium-dependent ATPases. Moreover...

β integrin tyrosine phosphorylation is a conserved mechanism for regulating talin-induced integrin activation; beta integrin tyrosine phosphorylation is a conserved mechanism for regulating talin-induced integrin activation

Anthis, N.; Haling, J.; Oxley, C.; Memo, M.; Wegener, K.; Lim, C.; Ginsberg, M.; Campbell, I.
Fonte: Amer Soc Biochemistry Molecular Biology Inc Publicador: Amer Soc Biochemistry Molecular Biology Inc
Tipo: Artigo de Revista Científica
Publicado em //2009 EN
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36.54%
Integrins are large membrane-spanning receptors fundamental to cell adhesion and migration. Integrin adhesiveness for the extracellular matrix is activated by the cytoskeletal protein talin via direct binding of its phosphotyrosine-binding-like F3 domain to the cytoplasmic tail of the β integrin subunit. The phosphotyrosine-binding domain of the signaling protein Dok1, on the other hand, has an inactivating effect on integrins, a phenomenon that is modulated by integrin tyrosine phosphorylation. Using full-length tyrosine-phosphorylated 15N-labeled β3, β1A, and β7 integrin tails and an NMR-based protein-protein interaction assay, we show that talin1 binds to the NPXY motif and the membrane-proximal portion of β3, β1A, and β7 tails, and that the affinity of this interaction is decreased by integrin tyrosine phosphorylation. Dok1 only interacts weakly with unphosphorylated tails, but its affinity is greatly increased by integrin tyrosine phosphorylation. The Dok1 interaction remains restricted to the integrin NPXY region, thus phosphorylation inhibits integrin activation by increasing the affinity of β integrin tails for a talin competitor that does not form activating membrane-proximal interactions with the integrin. Key residues governing these specificities were identified by detailed structural analysis...

Fragmentations of [M − H]⁻ anions of peptides containing tyrosine sulfate. Does the sulfate group rearrange? A joint experimental and theoretical study; Fragmentations of [M - H](-) anions of peptides containing tyrosine sulfate. Does the sulfate group rearrange? A joint experimental and theoretical study

Tran, T.; Wang, T.; Hack, S.; Bowie, J.
Fonte: John Wiley & Sons Ltd Publicador: John Wiley & Sons Ltd
Tipo: Artigo de Revista Científica
Publicado em //2013 EN
Relevância na Pesquisa
36.52%
RATIONALE: To investigate the fragmentations in the negative-ion electrospray mass spectra of peptides containing tyrosine sulfate. METHODS: Possible fragmentation mechanisms were explored using a Waters QTOF2 tandem mass spectrometer in concert with calculations at the CAM-B3LYP/6-311++g(d,p) level of theory. RESULTS: The major negative ion formed in the ESI-MS of peptides containing tyrosine sulfate is [(M-H)-SO3](-) and this process normally yields the base peak of the spectrum. The basic backbone cleavages of [(M-H)-SO3](-) allowed the sequence of the peptide to be determined. Rearrangement reactions involving the formation of HOSO3(-) and [(M-H)-H2SO4](-) yielded minor peaks with relative abundances ≤ 10% and ≤ 2%, respectively. CONCLUSIONS: The mass spectra of the [M-H](-) and [(M-H)-SO3](-) anions of peptides containing tyrosine sulfate allowed the position of the tyrosine sulfate group to be determined, together with the amino acid sequence of the peptide.; T. T. Nha Tran, Tianfang Wang, Sandra Hack and John H. Bowie

Protein Tyrosine Phosphatase Pez : its role in the regulation of cell-cell adhesions.

Wadham, Carol
Fonte: Universidade de Adelaide Publicador: Universidade de Adelaide
Tipo: Tese de Doutorado
Publicado em //2003
Relevância na Pesquisa
36.52%
The balance of tyrosine phosphorylation in the cell is maintained by the opposing actions of protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). Investigation into tyrosine phosphorylation was initially focused on the action of PTKs. However, research over the past decade has revealed that PTPs also play a key role in signal transduction. The multi-protein complexes that constitute the cell-cell adhesions in endothelial and epithelial tissues are dynamically restructured in response to extracellular and intracellular signalling. Tyrosine phosphorylation is involved in the regulation of both adherens junctions and tight junctions. Inhibitors of PTPs have been shown to disrupt cell-cell adhesions indicating that PTPs are important in maintaining adhesion integrity. The maintenance of a selectively permeable barrier is an essential function of endothelial cells, which are the cells that line the lumen of blood vessels. Therefore, it is important to understand the normal functioning of the proteins in the cell-cell adhesion complexes. The aims of this research were to ascertain which members of the PTP family are expressed in human umbilical vein endothelial cells (HUVEC) and to characterise a PTP that may potentially be involved in the regulation of cell-cell adhesions. A homology screen identified a cytosolic phosphatase...

The role of bacterial protein tyrosine phosphatases in the regulation of the biosynthesis of secreted polysaccharides

Standish, A.; Morona, R.
Fonte: Mary Ann Liebert Publicador: Mary Ann Liebert
Tipo: Artigo de Revista Científica
Publicado em //2014 EN
Relevância na Pesquisa
36.52%
SIGNIFICANCE: Tyrosine phosphorylation and associated protein tyrosine phosphatases are gaining prominence as critical mechanisms in the regulation of fundamental processes in a wide variety of bacteria. In particular, these phosphatases have been associated with the control of the biosynthesis of capsular polysaccharides and extracellular polysaccharides, critically important virulence factors for bacteria.RECENT ADVANCES: Deletion and over-expression of the phosphatases result in altered polysaccharide biosynthesis in a range of bacteria. The recent structures of associated auto-phosphorylating tyrosine kinases has suggested that the phosphatases may be critical for the cycling of the kinases between monomers and higher order oligomers. CRITICAL ISSUES: Additional substrates of the phosphatases apart from cognate kinases are currently being identified. These are likely to be critical to our understanding of the mechanism by which polysaccharide biosynthesis is regulated. FUTURE DIRECTIONS: Ultimately, these protein tyrosine phosphatases are an attractive target for the development of novel anti-microbials. This is particularly the case for the polymerase and histidinol phosphatase family, which are predominantly found in bacteria. Furthermore...

Tyrosine phosphorylation enhances activity of pneumococcal autolysin LytA

Standish, A.J.; Whittall, J.J.; Morona, R.
Fonte: Society for General Microbiology Publicador: Society for General Microbiology
Tipo: Artigo de Revista Científica
Publicado em //2014 EN
Relevância na Pesquisa
36.57%
For a long time tyrosine phosphorylation has been recognized as a crucial post translational regulatory mechanism in eukaryotes. However, only in the past decade has recognition been given to the crucial importance of bacterial tyrosine phosphorylation as an important regulatory feature of pathogenesis. This study describes the effect of tyrosine phosphorylation on the activity of a major virulence factor of the pneumococcus, the autolysin LytA, and a possible connection to the Streptococcus pneumoniae capsule synthesis regulatory proteins (CpsB, CpsC & CpsD). We show that in vitro pneumococcal tyrosine kinase, CpsD, and the protein tyrosine phosphatase, CpsB, act to phosphorylate and dephosphorylate LytA. Furthermore, this modulates LytA function in vitro with phosphorylated LytA binding more strongly to the choline analogue DEAE. A phospho-mimetic (Y264E) mutation of the LytA phosphorylation site displayed similar phenotypes as well as an enhanced dimerization capacity. Similarly, tyrosine phosphorylation increased LytA amidase activity, as evidenced by a turbidometric amidase activity assay. Similarly, when the phospho-mimetic mutation was introduced in the chromosomal lytA of S. pneumoniae, autolysis occurred earlier and at an enhanced rate. This study thus describes to our knowledge the first functional regulatory effect of tyrosine phosphorylation on a non-capsule related protein in the pneumococcus...

A light- and electron microscopic study of tyrosine

Tan, C.K.; Zhang, Y.L.; Wong, W. C.
Fonte: Murcia : F. Hernández Publicador: Murcia : F. Hernández
Tipo: Artigo de Revista Científica Formato: application/pdf
ENG
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36.54%
The present paper describes tyrosine hydroxylase-like immunoreactivity in the ciliary ganglion of monkey (Macaca fascicularis) and cat. Under the light microscope, in the monkey, about 7.6% of neurons were observed to be intensely stained, 27.7% moderately stained and 32.5% lightly stained. In the cat, 1.2% of neurons were intensely stained, 5.4% moderately stained and 10.1% lightly stained. Ultrastructurally, tyrosine hydroxylase-like immunoreactivity was observed in neurona1 somata, dendritic profiles and axons in both monkey and cat. Tyrosine hydroxylase-like immunoreactive dendritic profiles were synaptically contacted by tyrosine hydroxylase-negative axon terminals. In the monkey, tyrosine hydroxylase-like immunoreactive fibres were observed to enter the ciliary ganglion via the nasociliary nerve. Such fibres were observed to course among neurons within the ganglion and emerge in the short ciliary nerves. In contrast, tyrosine hydroxylase-like immunoreactive fibres were only occasionally observed in the cat.

Photo-oxidation of tyrosine in a bio-engineered bacterioferritin 'reaction centre'

Hingorani, Kastoori
Fonte: Universidade Nacional da Austrália Publicador: Universidade Nacional da Austrália
Tipo: Thesis (PhD)
EN_AU
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36.57%
The photosynthetic reaction centre (RC) is central to conversion of solar energy into chemical energy. In this thesis, in order to introduce the redox-active cofactors similar to that of the Photosystem II RC, a non-photosynthetic protein scaffold was used as an in vitro model. The protein tasked for this purpose was the heme containing bacterioferritin (BFR) protein found in E. coli. The BFR protein naturally expresses as a homodimer based on a 4-helix bundle monomer. Desirable properties included: (i) a promiscuos di-nuclear metal binding site which provides ligands for class II metals such as Mn (ii) a hydrophobic pocket at the dimer interface which can bind a photosensitive porphyrin, in this case a chlorin (Ce6), and (iii) presence of tyrosine residues proximal to the bound cofactors, which can be utilised as efficient electron-tunnelling intermediates. The work in this thesis extends earlier work in the group by refining and improving the BFR system. Several mutants were made and an improved protein expression system was developed. For these samples experiments demonstrated ligation of weakly coupled equivalent Mn2II,II at the di-nuclear binding site of apo-BFR, and binding of the photo-active pigment ZnCe6 in hydrophobic pocket of the protein. Light-induced electron transfer from proximal tyrosine residue(s) to the photo-oxidised ZnCe6+...

Structural and functional studies of bacterial protein tyrosine kinases

Lee, Daniel Cho-En
Fonte: Quens University Publicador: Quens University
Tipo: Tese de Doutorado Formato: 21191105 bytes; application/pdf
EN; EN
Relevância na Pesquisa
36.54%
While protein tyrosine kinases (PTKs) have been extensively characterized in eukaryotes, far less is known about their emerging counterparts in prokaryotes. Studies of close to 20 homologs of bacterial protein tyrosine (BY) kinases have inaugurated a blooming new field of research, all since just the end of the last decade. These kinases are key regulators in the polymerization and exportation of the virulence-determining polysaccharides which shield the bacterial from the non-specific defenses of the host. This research is aimed at furthering our understanding of the BY kinases through the use of X-ray crystallography and various in vitro and in vivo experiments. We reported the first crystal structure of a bacterial PTK, the C-terminal kinase domain of E. coli tyrosine kinase (Etk) at 2.5Å resolution. The fold of the Etk kinase domain differs markedly from that of eukaryotic PTKs. Based on the observed structure and supporting evidences, we proposed a unique activation mechanism for BY kinases in Gram-negative bacteria. The phosphorylation of tyrosine residue Y574 at the active site and the specific interaction of P-Y574 with a previously unidentified key arginine residue, R614, unblock the Etk active site and activate the kinase. Both in vitro kinase activity and in vivo antibiotics resistance studies utilizing structure-guided mutants further support the novel activation mechanism. In addition...

HAMSTER OVIDUCTIN ENHANCES TYROSINE PHOSPHORYLATION OF SPERM PROTEINS DURING CAPACITATION

Saccary, LAURELLE
Fonte: Quens University Publicador: Quens University
Tipo: Tese de Doutorado Formato: 8774028 bytes; application/pdf
EN; EN
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36.57%
Capacitation is essential for fertilization of ovulated oocytes. Capacitation is correlated with activation of a signal transduction pathway leading to protein tyrosine phosphorylation, an essential prerequisite for fertilization. Oviductin has been shown to bind to the acrosomal cap and the equatorial segment region of the sperm head. In light of findings reported in previous studies, we hypothesized that estrus stage-specific oviductin (EOV) enhances tyrosine phosphorylation. Immunofluorescent detection by light and confocal microscopy and immunogold labeling by electron microscopy and surface replica techniques were used to localize tyrosine phosphorylated proteins to the equatorial segment region and midpiece after incubation in medium in the presence or absence of EOV. In the presence of EOV, an increase in tyrosine phosphorylation in the equatorial segment region was observed as early as 5 minutes after incubation. On prolonging incubation in medium containing EOV immunostaining further increased, indicative of increased levels of tyrosine phosphorylation of sperm proteins as capacitation proceeds. Regardless of the presence or absence of EOV, phosphotyrosine expression was observed along the tail, specifically at the midpiece. However...

Effects of cigarette smoke on salivary protein tyrosine nitration

Weiner, D; Khankin, EV; Levy, Y; Aizenbud, D; Reznick, AZ
Fonte: BioMed Central Publicador: BioMed Central
Tipo: Artigo de Revista Científica
EN_US
Relevância na Pesquisa
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Introduction: Nitration of tyrosine and tyrosine-containing proteins and their roles in pathophysiology have just recently been reviewed. Despite low yields of tyrosine modifications, nitration of tyrosine residues may inactivate important proteins. Nitrotyrosine can be formed by various nitrating agents, including peroxynitrite. Thus, the occurrence of nitrotyrosine-containing proteins in vivo should be regarded as a general indication of tissue damage induced by reactive nitrogen species such as peroxynitrite. This strongly suggests that peroxynitrite could be formed in vivo under certain pathophysiological conditions. Objective: Our aim in this study was to elucidate the effect of cigarette smoke (CS) on nitrotyrosine formation in saliva proteins. Methods: We exposed saliva to CS, in vitro, and used Western Blotting (WB) and monoclonal anti-nitrotyrosine antibody to assess the level of saliva protein nitration. Results: As saliva contains extensive amounts of nitrites, it was no surprise that at basal levels, saliva proteins, albumin, and α-amylase all were already nitrated. The WB also revealed that with continuous exposure to CS the tyrosine nitration of both albumin and α-amylase is declining significantly after 3 h. A quite similar effect was seen after exposure to aldehydes...

PET with o-(2-18F-Fluoroethyl)-L-Tyrosine in Peripheral Tumors: First Clinical Results

Pauleit, Dirk; Stoffels, Gabriele; Schaden, Winfried; Hamacher, Kurt; Bauer, Dagmar; Tellmann, Lutz; Herzog, Hans; Broer, Stefan; Coenen, Heinz; Langen, Karl-Josef
Fonte: Society of Nuclear Medicine Publicador: Society of Nuclear Medicine
Tipo: Artigo de Revista Científica
Relevância na Pesquisa
36.52%
O-(2-18F-Fluoroethyl)-L-Tyrosine (18F-FET) PET has shown promising results in brain tumor diagnosis. The aim of this prospective study was to evaluate 18F-FET PET in comparison with 18F-FDG PET in patients with peripheral tumors. Methods: Forty-four consecutive patients with suspected malignant tumors underwent 18F-FET PET and 18F-FDG PET within 7 d. Whole-body PET studies were performed 1 h after intravenous injection of 370 MBq of 18F-FET or 18F-FDG. Six patients were excluded from the analysis because a malignant tumor could not be verified. In 38 patients (7 with colorectal cancer, 6 with pancreatic cancer, 9 with head-neck cancer, 4 with lymphomas, 3 with lung cancer, 3 with ovarian cancer, 4 with breast cancer, and 2 with prostatic cancer), 18F-FET PET and 18F-FDG PET were compared. Results: 18F-FET was positive in only 13 of 38 patients (8 with head-neck cancer, 3 with breast cancer, and 2 with lung cancer), whereas 18F-FDG exhibited increased uptake in 37 of 38 patients. All squamous cell carcinomas were found to be 18F-FET-positive tumors (8 head-neck cancer and 2 lung cancer), whereas most adenocarcinomas were found to be 18F-FET-negative tumors. In patients with colorectal cancer, pancreatic cancer, ovarian cancer, prostatic cancer...