54 research outputs found
Functional characterization of the evolutionarily divergent fern plastocyanin
De functie van eiwitten die betrokken zijn bij insulinewerking en glucose hemeostrase en de rol van genetische varianten hierin bij de ontwikkeling van type 2 diabete
Initial characteristics of RbcX proteins from Arabidopsis thaliana
Form I of Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase) is composed of eight large (RbcL) and eight small (RbcS) subunits. Assembly of these subunits into a functional holoenzyme requires the assistance of additional assembly factors. One such factor is RbcX, which has been demonstrated to act as a chaperone in the assembly of most cyanobacterial Rubisco complexes expressed in heterologous system established in Escherichia coli cells. Analysis of Arabidopsis thaliana genomic sequence revealed the presence of two genes encoding putative homologues of cyanobacterial RbcX protein: AtRbcX1 (At4G04330) and AtRbcX2 (At5G19855). In general, both RbcX homologues seem to have the same function which is chaperone activity during Rubisco biogenesis. However, detailed analysis revealed slight differences between them. AtRbcX2 is localized in the stromal fraction of chloroplasts whereas AtRbcX1 was found in the insoluble fraction corresponding with thylakoid membranes. Search for putative “partners” using mass spectrometry analysis suggested that apart from binding to RbcL, AtRbcX1 may also interact with β subunit of chloroplast ATP synthase. Quantitative RT-PCR analysis of AtRbcX1 and AtRbcX2 expression under various stress conditions indicated that AtRbcX2 is transcribed at a relatively stable level, while the transcription level of AtRbcX1 varies significantly. In addition, we present the attempts to elucidate the secondary structure of AtRbcX proteins using CD spectroscopy. Presented results are the first known approach to elucidate the role of RbcX proteins in Rubisco assembly in higher plants
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The allosteric regulation of axial/rhombic population in a "type 1" copper site: Multi-edge x-ray absorption spectroscopic and density functional Studies of Pseudoazurin
The co-existence of "axial" and "rhombic" coordination environments has been demonstrated in a "Type 1" copper site of Pseudoazurin. This observation opens up previously not considered interpretations for the relationship between geometry and electronic structure of the four coordinate copper site. The Met16 variants of pseudoazurin were considered as model systems for investigating the effect of weak interactions from the second coordination sphere. The correlation between geometric and electronic structures of "Type 1" copper site was evaluated by the multi-edge (Cu K-edge and S K-edge) X-ray absorption spectroscopy (XAS) of Met16 variants of pseudoazurin. The co-existing axial and rhombic sites in pseudoazurin were characterized by Culigand distances, effective nuclear charge, and Cu-S( Cys) covalency from XAS. The XAS results were correlated with DFT calculations for investigating the effect of protein environment from the inner-sphere and beyond around the Cu site. The combined experimental and theoretical results support the presence of a close correlation between outer sphere environment and inner sphere coordination environment. This is achieved in pseudoazurin by a previously undisclosed allosteric effect that involves a rearrangement of the protein tertiary structure
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The allosteric regulation of axial/rhombic population in a "type 1" copper site: Multi-edge x-ray absorption spectroscopic and density functional Studies of Pseudoazurin
The co-existence of "axial" and "rhombic" coordination environments has been demonstrated in a "Type 1" copper site of Pseudoazurin. This observation opens up previously not considered interpretations for the relationship between geometry and electronic structure of the four coordinate copper site. The Met16 variants of pseudoazurin were considered as model systems for investigating the effect of weak interactions from the second coordination sphere. The correlation between geometric and electronic structures of "Type 1" copper site was evaluated by the multi-edge (Cu K-edge and S K-edge) X-ray absorption spectroscopy (XAS) of Met16 variants of pseudoazurin. The co-existing axial and rhombic sites in pseudoazurin were characterized by Culigand distances, effective nuclear charge, and Cu-S( Cys) covalency from XAS. The XAS results were correlated with DFT calculations for investigating the effect of protein environment from the inner-sphere and beyond around the Cu site. The combined experimental and theoretical results support the presence of a close correlation between outer sphere environment and inner sphere coordination environment. This is achieved in pseudoazurin by a previously undisclosed allosteric effect that involves a rearrangement of the protein tertiary structure
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