scholarly journals Protein storage vacuoles form de novo during pea cotyledon development

1995 ◽  
Vol 108 (1) ◽  
pp. 299-310 ◽  
Author(s):  
B. Hoh ◽  
G. Hinz ◽  
B.K. Jeong ◽  
D.G. Robinson
Keyword(s):  
Membrane Protein ◽  
De Novo ◽  
Storage Proteins ◽  
Protein Body ◽  
Membrane System ◽  
Intermediate Stage ◽  
Integral Membrane Protein ◽  
Protein Storage ◽  
Protein Storage Vacuoles ◽  
Protein Storage Vacuole

We have investigated the formation of protein storage vacuoles in peas (Pisum sativum L.) in order to determine whether this organelle arises de novo during cotyledon development. A comparison of different stages in cotyledon development indicates that soluble protease activities decline and the amounts of storage proteins and the integral membrane protein of the protein body, alpha-TIP, increase during seed maturation. On linear sucrose density gradients we have been able to distinguish between two separate vesicle populations: one enriched in alpha-TIP, and one in TIP-Ma 27, a membrane protein characteristic of vegetative vacuoles. Both vesicle populations possess, however, PPase and V-ATPase activities. Conventionally fixed cotyledonary tissue at an intermediate stage in cotyledon development reveals the presence of a complex tubular-cisternal membrane system that seems to surround the pre-existing vacuoles. The latter gradually become compressed as a result of dilation of the former membrane system. This was confirmed immunocytochemically with the TIP-Ma 27 antiserum. Deposits of the storage proteins vicilin and legumin in the lumen, and the presence of alpha-TIP in the membranes of the expanding membrane system provide evidence of its identity as a precursor to the protein storage vacuole.

scholarly journals The protein storage vacuole

2001 ◽  
Vol 155 (6) ◽  
pp. 991-1002 ◽  
Author(s):  
Liwen Jiang ◽  
Thomas E. Phillips ◽  
Christopher A. Hamm ◽  
Yolanda M. Drozdowicz ◽  
Philip A. Rea ◽  
...  
Keyword(s):  
Seed Development ◽  
Secretory Pathway ◽  
Storage Proteins ◽  
Plant Seed ◽  
Protein Storage ◽  
Protein Storage Vacuoles ◽  
Protein Storage Vacuole ◽  
Membrane Bound ◽  
Lytic Vacuole ◽  
Storage Vacuole

Storage proteins are deposited into protein storage vacuoles (PSVs) during plant seed development and maturation and stably accumulate to high levels; subsequently, during germination the storage proteins are rapidly degraded to provide nutrients for use by the embryo. Here, we show that a PSV has within it a membrane-bound compartment containing crystals of phytic acid and proteins that are characteristic of a lytic vacuole. This compound organization, a vacuole within a vacuole whereby storage functions are separated from lytic functions, has not been described previously for organelles within the secretory pathway of eukaryotic cells. The partitioning of storage and lytic functions within the same vacuole may reflect the need to keep the functions separate during seed development and maturation and yet provide a ready source of digestive enzymes to initiate degradative processes early in germination.

scholarly journals A Novel Membrane Protein That Is Transported to Protein Storage Vacuoles via Precursor-Accumulating Vesicles

2001 ◽  
Vol 13 (10) ◽  
pp. 2361-2372 ◽  
Author(s):  
Naoto Mitsuhashi ◽  
Yasuko Hayashi ◽  
Yasuko Koumoto ◽  
Tomoo Shimada ◽  
Tomoko Fukasawa-Akada ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Protein Storage ◽  
Protein Storage Vacuoles

scholarly journals Insights into the Relationship between Cobamide Synthase and the Cell Membrane

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Victoria L. Jeter ◽  
Jorge C. Escalante-Semerena
Keyword(s):  
Cell Membrane ◽  
Membrane Protein ◽  
Lipid Bilayer ◽  
De Novo ◽  
Integral Membrane Protein ◽  
Phospholipid Bilayer ◽  
Multienzyme Complex ◽  
Coenzyme B ◽  
Lower Ligand

ABSTRACT Cobamides are cobalt-containing cyclic tetrapyrroles used by cells from all domains of life but only produced de novo by some bacteria and archaea. The “late steps” of the adenosylcobamide biosynthetic pathway are responsible for the assembly of the nucleotide loop and are required during de novo synthesis and precursor salvaging. These steps are characterized by activation of the corrin ring and lower ligand base, condensation of the activated precursors to adenosylcobamide phosphate, and removal of the phosphate, yielding a complete adenosylcobamide molecule. The condensation of the activated corrin ring and lower ligand base is performed by an integral membrane protein, cobamide (5′ phosphate) synthase (CobS), and represents an important convergence of two pathways necessary for nucleotide loop assembly. Interestingly, membrane association of this penultimate step is conserved among all cobamide producers, yet the physiological relevance of this association is not known. Here, we present the purification and biochemical characterization of the CobS enzyme of the enterobacterium Salmonella enterica subsp. enterica serovar Typhimurium strain LT2, investigate its association with liposomes, and quantify the effect of the lipid bilayer on its enzymatic activity and substrate affinity. We report a purification scheme that yields pure CobS protein, allowing in vitro functional analysis. Additionally, we report a method for liposome reconstitution of CobS, allowing for physiologically relevant studies of this inner membrane protein in a phospholipid bilayer. In vitro and in vivo data reported here expand our understanding of CobS and the implications of membrane-associated adenosylcobamide biosynthesis. IMPORTANCE Salmonella is a human pathogen of worldwide importance, and coenzyme B12 is critical for the pathogenic lifestyle of this bacterium. The importance of the work reported here lies on the improvements to the methodology used to isolate cobamide synthase, a polytopic integral membrane protein that catalyzes the penultimate step of coenzyme B12 biosynthesis. This advance is an important step in the analysis of the proposed multienzyme complex responsible for the assembly of the nucleotide loop during de novo coenzyme B12 biosynthesis and for the assimilation of incomplete corrinoids from the environment. We proposed that cobamide synthase is likely localized to the cell membrane of every coenzyme B12-producing bacterium and archaeum sequenced to date. The new knowledge of cobamide synthase advances our understanding of the functionality of the enzyme in the context of the lipid bilayer and sets the foundation for the functional-structural analysis of the aforementioned multienzyme complex.

scholarly journals Staphylococcus aureus HemX Modulates Glutamyl-tRNA Reductase Abundance To Regulate Heme Biosynthesis

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Jacob E. Choby ◽  
Caroline M. Grunenwald ◽  
Arianna I. Celis ◽  
Svetlana Y. Gerdes ◽  
Jennifer L. DuBois ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Membrane Protein ◽  
De Novo ◽  
Common Factor ◽  
Integral Membrane Protein ◽  
Heme Biosynthesis ◽  
Bacterial Physiology ◽  
Heme Synthesis ◽  
Regulatory Processes ◽  
High Concentrations

ABSTRACTStaphylococcus aureusis responsible for a significant amount of devastating disease. Its ability to colonize the host and cause infection is supported by a variety of proteins that are dependent on the cofactor heme. Heme is a porphyrin used broadly across kingdoms and is synthesizedde novofrom common cellular precursors and iron. While heme is critical to bacterial physiology, it is also toxic in high concentrations, requiring that organisms encode regulatory processes to control heme homeostasis. In this work, we describe a posttranscriptional regulatory strategy inS. aureusheme biosynthesis. The first committed enzyme in theS. aureusheme biosynthetic pathway, glutamyl-tRNA reductase (GtrR), is regulated by heme abundance and the integral membrane protein HemX. GtrR abundance increases dramatically in response to heme deficiency, suggesting a mechanism by whichS. aureusresponds to the need to increase heme synthesis. Additionally, HemX is required to maintain low levels of GtrR in heme-proficient cells, and inactivation ofhemXleads to increased heme synthesis. Excess heme synthesis in a ΔhemXmutant activates the staphylococcal heme stress response, suggesting that regulation of heme synthesis is critical to reduce self-imposed heme toxicity. Analysis of diverse organisms indicates that HemX is widely conserved among heme-synthesizing bacteria, suggesting that HemX is a common factor involved in the regulation of GtrR abundance. Together, this work demonstrates thatS. aureusregulates heme synthesis by modulating GtrR abundance in response to heme deficiency and through the activity of the broadly conserved HemX.IMPORTANCEStaphylococcus aureusis a leading cause of skin and soft tissue infections, endocarditis, bacteremia, and osteomyelitis, making it a critical health care concern. Development of new antimicrobials againstS. aureusrequires knowledge of the physiology that supports this organism’s pathogenesis. One component of staphylococcal physiology that contributes to growth and virulence is heme. Heme is a widely utilized cofactor that enables diverse chemical reactions across many enzyme families.S. aureusrelies on many critical heme-dependent proteins and is sensitive to excess heme toxicity, suggestingS. aureusmust maintain proper intracellular heme homeostasis. BecauseS. aureusprovides heme for heme-dependent enzymes via synthesis from common precursors, we hypothesized that regulation of heme synthesis is one mechanism to maintain heme homeostasis. In this study, we identify thatS. aureusposttranscriptionally regulates heme synthesis by restraining abundance of the first heme biosynthetic enzyme, GtrR, via heme and the broadly conserved membrane protein HemX.

One Vacuole or two Vacuoles: Do Protein Storage Vacuoles Arise de novo during Pea Cotyledon Development?

1995 ◽  
Vol 145 (5-6) ◽  
pp. 654-664 ◽  
Author(s):  
David G. Robinson ◽  
Birgit Hoh ◽  
Giselbert Hinz ◽  
Byung-Kap Jeong
Keyword(s):  
De Novo ◽  
Protein Storage ◽  
Protein Storage Vacuoles

scholarly journals Vacuolar Storage Proteins Are Sorted in the Cis-Cisternae of the Pea Cotyledon Golgi Apparatus

2001 ◽  
Vol 152 (1) ◽  
pp. 41-50 ◽  
Author(s):  
Stefan Hillmer ◽  
Ali Movafeghi ◽  
David G. Robinson ◽  
Giselbert Hinz
Keyword(s):  
Golgi Apparatus ◽  
Storage Proteins ◽  
Spatial Separation ◽  
Golgi Stack ◽  
Protein Storage ◽  
Protein Storage Vacuole ◽  
Vacuolar Sorting ◽  
Sorting Receptor ◽  
Storage Vacuole ◽  
Dense Vesicles

Developing pea cotyledons contain functionally different vacuoles, a protein storage vacuole and a lytic vacuole. Lumenal as well as membrane proteins of the protein storage vacuole exit the Golgi apparatus in dense vesicles rather than in clathrin-coated vesicles (CCVs). Although the sorting receptor for vacuolar hydrolases BP-80 is present in CCVs, it is not detectable in dense vesicles. To localize these different vacuolar sorting events in the Golgi, we have compared the distribution of vacuolar storage proteins and of α-TIP, a membrane protein of the protein storage vacuole, with the distribution of the vacuolar sorting receptor BP-80 across the Golgi stack. Analysis of immunogold labeling from cryosections and from high pressure frozen samples has revealed a steep gradient in the distribution of the storage proteins within the Golgi stack. Intense labeling for storage proteins was registered for the cis-cisternae, contrasting with very low labeling for these antigens in the trans-cisternae. The distribution of BP-80 was the reverse, showing a peak in the trans-Golgi network with very low labeling of the cis-cisternae. These results indicate a spatial separation of different vacuolar sorting events in the Golgi apparatus of developing pea cotyledons.

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