RESUMO
Endoplasmic reticulum (ER)-derived compartments are found in many plant species. Although it has been assumed that aggregation induces formation of the ER-derived compartments in plant seed cells, the effect of aggregation on the trafficking from the ER to the Golgi has not yet been elucidated. In this study, we used an aggregated type of red fluorescent protein (DsRED) to investigate the effect of aggregation on sorting in seed cells. DsRED fused to the Golgi sorting determinant was found mainly in large globular structures derived from the ER where ER-resident proteins were excluded. These results indicate that aggregation of the Golgi protein blocks transport from the ER to the Golgi.
Assuntos
Retículo Endoplasmático/metabolismo , Complexo de Golgi/metabolismo , Proteínas de Plantas/metabolismo , Animais , Antozoários , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/ultraestrutura , Brefeldina A/farmacologia , Retículo Endoplasmático/efeitos dos fármacos , Complexo de Golgi/efeitos dos fármacos , Proteínas Luminescentes/química , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Mutação , Pentosiltransferases/química , Pentosiltransferases/genética , Pentosiltransferases/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Plantas Geneticamente Modificadas/ultraestrutura , Conformação Proteica , Dobramento de Proteína , Transporte Proteico , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Sementes/metabolismo , Sementes/ultraestrutura , Proteína Vermelha FluorescenteRESUMO
beta-Conglycinin is composed of three kinds of subunit: alpha, alpha' and beta. A phagocytosis-stimulating peptide sequence (MITLAIPVNKPGR), soymetide, exists in the alpha' subunit of beta-conglycinin. Met at N terminus of the soymetide is essential for the activity. When Thr at the third residue from N terminus of the soymetide is replaced by Phe or Trp, the phagocytosis-stimulating activity greatly increases (ThrAssuntos
Globulinas/química
, Glycine max/química
, Proteínas de Soja/química
, Antígenos de Plantas
, Varredura Diferencial de Calorimetria
, Dicroísmo Circular
, Globulinas/biossíntese
, Globulinas/genética
, Temperatura Alta
, Modelos Moleculares
, Fagocitose/fisiologia
, Estrutura Terciária de Proteína
, Proteínas de Armazenamento de Sementes
, Proteínas de Soja/biossíntese
, Proteínas de Soja/genética
, Glycine max/genética
RESUMO
A patient with cholesteryl ester transfer protein (CETP) deficiency presents with marked hyperalphalipoproteinemia (HALP). To investigate the contribution of CETP deficiency to the cause of HALP (HDL-C> or =1.94 mmol/l, 75 mg/dl), we investigated the CETP activities and the prevalence of genetic CETP mutations among 624 Japanese HALP subjects. The subjects were screened for four known genetic CETP mutations (intron 14 splicing defect (In14), exon 15 missense mutation (Ex15), intron 10 splicing defect (In10) and exon 6 nonsense mutation (Ex6)). We found the frequency of the patients with reduced CETP activity (<75% of normal controls) to be 55.5 and 64.1% in a high HDL group (1.94< or =HDL-C<2.59 mmol/l) and a marked HALP group (HDL-C> or =2.59 mmol/l, 100 mg/dl), respectively. At least one of the four mutations was identified in 65.7% of subjects with reduced CETP activities and 57.5% of subjects with marked HALP. The In14 and Ex15 mutations were very common in HALP subjects and the frequency of In10 mutation and Ex6 mutation was quite low. To investigate the impact of genetic CETP mutation on the phenotypes, we compared the plasma lipid levels and CETP activities between the subjects with two common mutations. All In14 homozygotes showed marked HALP, while marked HALP is less frequent (64.3%) in Ex15 homozygotes. HDL-C levels in Ex15 heterozygotes were significantly higher than those of In14 heterozygotes, suggesting the mutation has dominant negative effects on CETP activity in vivo. Some cases with In14 (5.7%) or Ex15 (7.2%) mutation showed low HDL-C levels. We conclude that CETP deficiency is a major cause of HALP; nevertheless CETP deficiency is not necessarily HALP.
Assuntos
Apolipoproteínas/sangue , Proteínas de Transporte/genética , Colesterol/sangue , Glicoproteínas , Hiperlipoproteinemias/genética , Adulto , Idoso , Proteínas de Transporte/metabolismo , Proteínas de Transferência de Ésteres de Colesterol , Feminino , Humanos , Hiperlipoproteinemias/epidemiologia , Japão/epidemiologia , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Fenótipo , PrevalênciaRESUMO
The constituent subunits of seed storage protein 11S globulin have two disulfide bonds that are common among 11S globulins from legume and nonlegume seeds. In the case of the A1aB1b subunit of soybean 11S globulin, glycinin, Cys12-Cys45 and Cys88-Cys298 are observed by X-ray crystallography. The significance of these two disulfide bonds for structural stability was investigated by mutagenesis of Cys12 to Gly and of Cys88 to Ser. The disulfide bond-deficient mutants C12G and C88S could form the correct conformations identical to that of the wild-type proglycinin except in the vicinities of the mutation sites C12 and C88 as shown by their crystal structures. Thermal stability monitored by differential scanning calorimetry of the mutants indicated that the contribution of these disulfide bonds to the thermal stability of proglycinin A1aB1b is low, although there is a small difference in the extent of the contribution between the two disulfide bonds (Cys12-Cys45 > Cys88-Cys298). The contribution of Cys88-Cys298 to the resistance of proglycinin A1aB1b to proteinase digestion is higher than that of Cys12-Cys45. Possible effects of structure on the different properties of C12G and C88S are discussed.