Dataset Information

Phosphorylation of purified recombinant mouse Ric-8B

ABSTRACT: Analysis of recombinant mouse Ric-8B protein phosphorylation. Ric-8B was purified from insect cells and E.coli and both treated with protein kinase CK2 and re-purified prior to LC-MS/MS. Another Ric-8B sample from insect cells was not treated with CK2 and also subjected to LC-MS/MS.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Frank Kwarcinski

LAB HEAD: Gregory G. Tall

PROVIDER: PXD036645 | Pride | 2024-05-23

REPOSITORIES: Pride

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Publications

Structures of Ric-8B in complex with Gα protein folding clients reveal isoform specificity mechanisms.

Papasergi-Scott Makaía M MM Kwarcinski Frank E FE Yu Maiya M Panova Ouliana O Ovrutsky Ann M AM Skiniotis Georgios G Tall Gregory G GG

Structure (London, England : 1993) 20230316 5

Mammalian Ric-8 proteins act as chaperones to regulate the cellular abundance of heterotrimeric G protein α subunits. The Ric-8A isoform chaperones Gαi/o, Gα12/13, and Gαq/11 subunits, while Ric-8B acts on Gαs/olf subunits. Here, we determined cryoelectron microscopy (cryo-EM) structures of Ric-8B in complex with Gαs and Gαolf, revealing isoform differences in the relative positioning and contacts between the C-terminal α5 helix of Gα within the concave pocket formed by Ric-8 α-helical repeat el ...[more]

PMID: 36931277

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Project description:Background: Lignocellulosic biomass is a promising renewable feedstock for the microbial production of fuels. To release the major fermentable sugars such as glucose and xylose, pretreatment and enzymatic hydrolysis of biomass feedstock are needed. During this process, many toxic compounds are produced or introduced which subsequently inhibit microbial growth and eventually the production rate and yield. Acetate is one of the major inhibitors liberated from hemicelluloses during dilute acid pretreatment. An understanding of the toxic effects of acetate on the fermentation microorganism is critical to improving biofuel yields in the process. In addition, the efficient utilization of mixed sugars of glucose and xylose in the presence of hydrolysate inhibitors is crucial for economic biofuel production. Results: We have observed previously that some pretreatment inhibitors affect growth and performance in Zymomonas mobilis 8b differently when different sugars (e.g. glucose or xylose) are used as substrate. To investigate this phenomenon at the cellular level, microarray technology was used to investigate the acetate stress responses of Z. mobilis 8b when using single carbon sources of glucose or xylose, and mixed sugars of glucose and xylose. We designed a microarray based on the most up-to-date genome annotation for both coding sequences and intergenic regions. In the presence of acetate, 8b still can utilize all the glucose (though xylose utilization was inhibited) with similar ethanol yield although the growth, final biomass, and ethanol production rate were reduced. The presence of acetate caused genes related to biosynthesis, flagellar system, and glycolysis to be downregulated, and genes related to stress responses and energy metabolism to be upregulated. Our result indicates that Z. mobilis utilized different mechanism for xylose utilization compared to that of glucose, with even more dramatic results than those caused by treatment of the culture with the inhibitor acetate. Our study also suggests that redox imbalance caused by stressful conditions may trigger a metabolic reaction that leads to the accumulation of toxic intermediates such as xylitol, but Z. mobilis appears to be capable of managing its carbon and energy metabolism through the control of individual reactions to overcome the inhibition caused by stressful conditions. Several target gene candidates based on transcriptomic result have been selected for genetic manipulation and a TonB-dependent receptor knockout mutant was confirmed to have advantage on acetate tolerance. Conclusions: We have gained insights into the molecular responses of the model ethanologenic bacterium Z. mobilis to the inhibitor acetate when grown in different sugar sources. These insights will facilitate future metabolic modeling studies and help further strain metabolic engineering efforts for better xylose utilization and acetate tolerance. Two series of microarray studies using total RNA extracted from Zymomonas mobilis subsp mobilis 8b (an xylose-utilizing recombinant) were carried out to investigate the effect of carbon source and acetate on Z. mobilis. One study compared the acetate effect in either glucose or xylose at exponential phase and another study investigated the acetate effect in mixed sugar of glucose and xylose at three growth phases of exponential, transition, and stationary. Tthree biological replicates were used for each condition.

Dataset Information

Phosphorylation of purified recombinant mouse Ric-8B

Publications

Structures of Ric-8B in complex with Gα protein folding clients reveal isoform specificity mechanisms.

Similar Datasets

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