Assuntos
Povo Asiático/genética , Colágeno Tipo VII/genética , Epidermólise Bolhosa Distrófica/genética , Glicina/genética , Cloridrato de Olopatadina/administração & dosagem , Vitamina E/administração & dosagem , Administração Oral , Administração Tópica , Substituição de Aminoácidos/genética , Anti-Inflamatórios não Esteroides/administração & dosagem , Criança , Colágeno Tipo VII/química , Epidermólise Bolhosa Distrófica/diagnóstico , Epidermólise Bolhosa Distrófica/tratamento farmacológico , Glicina/química , Humanos , Masculino , Estrutura Secundária de Proteína , Resultado do TratamentoRESUMO
Trehalose dimycolate (TDM) is a major surface-exposed mycolyl glycolipid that contributes to the hydrophobic cell wall architecture of mycobacteria. Nevertheless, because of its potent adjuvant functions, pathogenic mycobacteria appear to have evolved an evasive maneuver to down-regulate TDM expression within the host. We have shown previously that Mycobacterium tuberculosis (M.tb) and Mycobacterium avium (M.av), replace TDM with glucose monomycolate (GMM) by borrowing host-derived glucose as an alternative substrate for the FbpA mycolyltransferase. Mycobacterium leprae (M.le), the causative microorganism of human leprosy, is also known to down-regulate TDM expression in infected tissues, but the function of its mycolyltransferases has been poorly analysed. We found that, unlike M.tb and M.av FbpA enzymes, M.av FbpA was unexpectedly inefficient in transferring alpha-branched mycolates, resulting in impaired production of both TDM and GMM. Molecular modelling and mutational analysis indicated that a bulky side chain of leucine at position 130 of M.le FbpA obstructed the intramolecular tunnel that was proposed to accommodate the alpha-branch portion of the substrates. Notably, even after a highly reductive evolution, M.le FbpA remained functional in terms of transferring unbranched acyl chains, suggesting a role that is distinct from that as a mycolyltransferase.
Assuntos
Aciltransferases/metabolismo , Glicolipídeos/metabolismo , Mycobacterium leprae/enzimologia , Ácidos Micólicos/metabolismo , Sequência de Aminoácidos , Substituição de Aminoácidos/genética , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Ácidos Graxos/metabolismo , Cinética , Leucina/metabolismo , Dados de Sequência Molecular , Proteínas Mutantes/metabolismo , Ligação Proteica , Especificidade por SubstratoRESUMO
Leprosy is the major cause of non-traumatic neuropathy. Herein, we investigated the role of ninjurin 1, an adhesion molecule involved in nerve regeneration in leprosy. Our results demonstrated that M. leprae stimulates in vitro up-regulation of ninjurin mRNA in cultured Schwann and blood cells as well as in vivo mRNA and protein expression in leprosy nerve biopsies. A polymorphism (asp110ala) was investigated in a case-control study (1123 individuals) and no association was found with leprosy per se or with disseminated forms. Nevertheless, ala110 was associated with functional nerve impairment (OR=2.42; p=0.02 for ala/ala) and with lower mRNA levels. Our data suggests that asp110ala could be a valuable genetic marker of nerve damage in leprosy.
Assuntos
Moléculas de Adesão Celular Neuronais/genética , Hanseníase/complicações , Hanseníase/genética , Fatores de Crescimento Neural/genética , Nervos Periféricos/metabolismo , Doenças do Sistema Nervoso Periférico/genética , Polimorfismo de Nucleotídeo Único/genética , Adulto , Alanina/genética , Substituição de Aminoácidos/genética , Ácido Aspártico/genética , Moléculas de Adesão Celular Neuronais/química , Análise Mutacional de DNA , Feminino , Marcadores Genéticos/genética , Predisposição Genética para Doença/genética , Testes Genéticos , Genótipo , Humanos , Imunidade Inata/genética , Hanseníase/fisiopatologia , Masculino , Pessoa de Meia-Idade , Fatores de Crescimento Neural/química , Nervos Periféricos/patologia , Nervos Periféricos/fisiopatologia , Doenças do Sistema Nervoso Periférico/diagnóstico , Doenças do Sistema Nervoso Periférico/fisiopatologia , RNA Mensageiro/metabolismo , Regulação para Cima/genéticaRESUMO
The present study reports, for the first time, that the recombinant hsp65 from Mycobacterium leprae (chaperonin 2) displays a proteolytic activity toward oligopeptides. The M. leprae hsp65 proteolytic activity revealed a trypsin-like specificity toward quenched fluorescence peptides derived from dynorphins. When other peptide substrates were used (beta-endorphin, neurotensin, and angiotensin I), the predominant peptide bond cleavages also involved basic amino acids in P(1), although, to a minor extent, the hydrolysis involving hydrophobic and neutral amino acids (G and F) was also observed. The amino acid sequence alignment of the M. leprae hsp65 with Escherichia coli HslVU protease suggested two putative threonine catalytic groups, one in the N-domain (T(136), K(168), and Y(264)) and the other in the C-domain (T(375), K(409), and S(502)). Mutagenesis studies showed that the replacement of K(409) by A caused a complete loss of the proteolytic activity, whereas the mutation of K(168) to A resulted in a 25% loss. These results strongly suggest that the amino acid residues T(375), K(409), and S(502) at the C-domain form the catalytic group that carries out the main proteolytic activity of the M. leprae hsp65. The possible pathophysiological implications of the proteolytic activity of the M. leprae hsp65 are now under investigation in our laboratory.