Granule-associated Proteins Research Articles

The role of starch granule-associated proteins in enhancing the strength of indica rice starch gels

The gel properties of indica rice starch strongly influence the quality of foods in which it is used. This study investigated the effects of chymotrypsin treatment on the rheological and pasting properties of starch prepared by different treatment methods. The results demonstrated that all the starches contained starch granule-associated proteins (SGAPs). Specifically, chymotrypsin-treated fermented extracted starch (E-FS) and chymotrypsin-treated alkali extracted starch (E-AS) exhibited a storage modulus (Gʹ) about 4 times higher than that of fermented extracted starch (FS) and alkali-extracted starch (AS). The flow consistency coefficients (K), loss moduli (Gʺ) and setback values tended to increase, while the characteristic rheological indices (n) and loss tangent (tan δ) tended to decrease after chymotrypsin treatment. This suggested that the starch gel was significantly enhanced by the proteinase modification. The protein composition of AS and E-AS only contained the 66 kDa of granule-bound starch synthase I (GBSS I) subunit. Therefore, the significant increase in gel strength was attributed to the exclusive modification effects of GBSS I. Microstructural analysis also confirmed that the three-dimensional gel network of indica rice starch was formed by a backbone of SGAP polymers filled with starch granules. The mechanism underlying GBSS I-enhanced gel was also elucidated based on the sequence and structure of GBSS I. The gel network formed by SGAP polymers of FS and AS was mainly caused by disulfide bonds between GBSS Is, while that of E-FS and E-AS was mainly caused by hydrophobic interactions. • The gel properties of indica starch treated by different methods were studied. • Enzymatic modification of starch granule-associated proteins (SGAPs) increased the starch gel strength significantly. • The SGAPs were mainly composed of granule-bound starch synthase I (GBSS I). • Gel formation driving force changed from disulfide bonds to hydrophobic interactions. • The gel structure comprised a polymerized SGAP network filled with starch granules.

Amphiphilic Cationic Peptide-Coated PHA Nanosphere as an Efficient Vector for Multiple-Drug Delivery.

Amphiphilic core–shell (ACS) nanoparticles are gaining increasing research interest for multi-drug delivery in cancer therapy. In this work, a new cationic peptide-coated PHA nanosphere was prepared by self-assembly of a hydrophobic core of biodegradable poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) and a hydrophilic shell of fusion proteins of PHA granule-associated protein (PhaP) and cationic peptide RALA through a strong hydrophobic effect. The hydrophobic drug curcumin (Cur) was encapsulated in PHBHHx nanoparticles. The chemotherapy drug 5-fluorouracil (5-FU) was administered in the form of its metabolite oligomeric 5-fluorodeoxyuridine (FUdR). Fifteen consecutive FUdR (FUdR15S) were adsorbed on the surface of PHBHHx nanoparticles by electrostatic interaction with RALA to form Cur@PHBX-PR/FUdR15S. Such amphiphilic cationic nanospheres had 88.3% EE of Cur and the drug loading of Cur and FUdR were 7.8% and 12.1%. The dual-drug-loaded nanospheres showed a time-differential release of Cur and FUdR. In addition, Cur@PHBX-PR/FUdR15S exhibited excellent anticancer activity and played a vital role in promoting the synergistic effect of FUdR and Cur in gastric cancer cells. The exploration of antitumor mechanisms demonstrated that Cur improved the activity of apoptosis-related proteins and cancer cells sensitized to FUdR. This amphiphilic core–shell system can serve as a general platform for sequential delivery of multiple drugs to treat several cancer cells.

Rhino-Orbital-Cerebral Mycosis and Extranodal Natural Killer or/and T-Cell Lymphoma, Nasal Type.

BackgroundExtranodal natural killer/T-cell lymphoma, nasal type is a syndrome of middle face destruction with an association to Epstein-Barr virus. Fungi have been recovered from the diseased tissue now and then but were often seen as a lymphoma-associated secondary infection. However, there are ENKTL-NT cases with the recoveries of fungi and complete recovery with antifungal therapy, which are quite similar to rhino-orbital-cerebral mycosis (ROCM) that often confuses the physicians.MethodsWe searched Medline for English-language manuscripts limited to “human” and “case reports,” “letters,” “reviews,” and “clinical conferences” from 1966 to 2022. We used MeSH terms “lymphoma, extranodal nk-t-cell” [MeSH Terms] or “lethal midline granuloma” [MeSH Terms], in combination with MeSH terms “microbiology” [subheading] or “microbiology” [all fields] or “fungi” [all fields] or “fungi” [MeSH Terms] for ENKTL-NT with infections. We used MeSH terms “Mycoses” in combination with “Nose” [Mesh] OR “Orbital Diseases” [Mesh] for rhino-orbital-cerebral fungal infections.ResultsWe appraised 149 included articles and extracted references related to ENKTL-NT and/or ROCM. Themes and subcategories were subsequently derived. Our findings revealed that ROCM and ENKTL-NT are characterized by progressive and destructive ulcers in the midline face or rhino-orbital structures. ROCM is mainly caused by fungi in the order of Mucorales, and ENKTL-NT is usually associated with Epstein-Barr virus and sometimes fungi. Radiologically, both are characterized by non-specific features of sinusitis, soft tissue infection, and necrosis. Pathologically, ROCM and ENKTL-NT share the same characteristics of inflammation, necrosis, and granuloma. ROCM is characterized by the detection of fungi in tissue, while ENKTL-NT is typically positive for NK/T-cell markers and cytotoxic granule-associated proteins, proliferation, and vascular damage of angioinvasion, which could be incited by Mucor irregularis and Rhizopus arrhizus in patients and mice.ConclusionENKTL-NT and ROCM share many similarities in clinical presentations, radiology, and histopathology, and might have the same etiology. This may explain why the two diseases are tangled together in the reported cases, and suggests the role that the fungi may play in the development of these ENKTL-NT/ROCM diseases. The reason why ENKTL-NT and ROCM are sometimes confused is that the main pathogens of ROCM, Mucor irregularis and Rhizopus arrhizus, are the fungal causative agents of ENKTL-NT.

Three Diverse Granule Preparation Methods for Proteomic Analysis of Mature Rice (Oryza sativa L.) Starch Grain

Starch is the primary form of reserve carbohydrate storage in plants. Rice (Oryza sativa L.) is a monocot whose reserve starch is organized into compounded structures within the amyloplast, rather than a simple starch grain (SG). The mechanism governing the assembly of the compound SG from polyhedral granules in apposition, however, remains unknown. To further characterize the proteome associated with these compounded structures, three distinct methods of starch granule preparation (dispersion, microsieve, and flotation) were performed. Phase separation of peptides (aqueous trypsin-shaving and isopropanol solubilization of residual peptides) isolated starch granule-associated proteins (SGAPs) from the distal proteome of the amyloplast and the proximal ‘amylome’ (the amyloplastic proteome), respectively. The term ‘distal proteome’ refers to SGAPs loosely tethered to the amyloplast, ones that can be rapidly proteolyzed, while proximal SGAPs are those found closer to the remnant amyloplast membrane fragments, perhaps embedded therein—ones that need isopropanol solvent to be removed from the mature organelle surface. These two rice starch-associated peptide samples were analyzed using nano-liquid chromatography–tandem mass spectrometry (Nano-HPLC-MS/MS). Known and novel proteins, as well as septum-like structure (SLS) proteins, in the mature rice SG were found. Data mining and gene ontology software were used to categorize these putative plastoskeletal components as a variety of structural elements, including actins, tubulins, tubulin-like proteins, and cementitious elements such as reticulata related-like (RER) proteins, tegument proteins, and lectins. Delineating the plastoskeletal proteome begins by understanding how each starch granule isolation procedure affects observed cytoplasmic and plastid proteins. The three methods described herein show how the technique used to isolate SGs differentially impacts the subsequent proteomic analysis and results obtained. It can thus be concluded that future investigations must make judicious decisions regarding the methodology used in extracting proteomic information from the compound starch granules being assessed, since different methods are shown to yield contrasting results herein. Data are available via ProteomeXchange with identifier PXD032314.

Nonstructural Protein 1 of SARS Coronavirus Interacts with Stress Granule Protein G3BP1 and Accumulates in the Stress Granule

The nonstructural protein 1 (nsp1) of severe acute respiratory syndrome coronavirus (SARS‐CoV and SARS‐CoV‐2) orchestrates a multi‐pronged mechanism to suppress host gene expression, also known as the host shutoff. The C‐terminus of nsp1 competes for the mRNA binding site in the 40S ribosome, preventing host translation from host mRNAs. Additionally, nsp1 triggers selective cleavage of host mRNAs while keeping the viral RNA intact. Nsp1 of SARS‐CoV and SARS‐CoV‐2 share 84% sequence identity, prompting us to believe they share a similar mechanism of host shutoff. So far identification of nsp1‐interacting partners has isolated stable complexes of nsp1 with the 40S and 60S ribosomal proteins. However, these complexes do not reveal any transient interaction nsp1 might have with other host proteins to suppress cellular function. Using proximity‐dependent biotinylation to capture the transient interactions of nsp1 with other cellular proteins, we found multiple subunits of the nuclear pre‐mRNA processing complex and cellular stress granule‐associated proteins. Here, we show that nsp1 interacts with the majority of these pre‐mRNA processing and stress granule‐associated proteins through its C‐terminal domain, the region responsible for binding the 40S ribosome. In addition, nsp1 co‐immunoprecipitates with stress granule‐associated protein G3BP1 and eIF4G. Upon induction of stress, these proteins accumulate in nonmembranous organelles called stress granules that are known to store stalled translation complexes. Using both immunofluorescence and immunoblot against stress granule‐associated proteins we show that nsp1 disrupts the maturation of stress granules.

Architecture of outer shell and inner blocklets of rice starch granule is related to starch granule-associated proteins

Waxy, low-amylose and high-amylose rice starches were chosen to separate inner blocklets and outer shell from their granules under incomplete gelatinization. Isolated outer shells and swollen or merged inner blocklets, based on morphological structure of broken granules, were used as a probe to study the mechanism of differences between outer and inner blocklets formation, through molecular characteristics and starch granule-associated proteins (SGAPs) analyses. On the basis of molecular composition, it was concluded that shorter branched-chain amylopectin and some amylose chains were involved in the formation of inner blocklets (shells). While the stronger interaction occurred among outer blocklets (shell), those are composed of longer branched-chains amylopectin and few amylose molecules. SGAPs types and content of inner shells (blocklets) were higher than outer shell (blocklets). SGAPs not only interacted with amylose chains but also with amylopectin chains. The enzymatic machinery for biosynthesis, such as starch branching enzyme, plays a critical role in blocklet architecture.

Repression of the stress granule protein G3BP2 inhibits immune checkpoint molecule PD-L1.

Mounting evidence suggests that cancer stemness and immunosuppression are related, but the underlying mechanisms behind these are not clear. We previously reported that the stress granule-associated protein G3BP2 is involved in the regulation of tumor-initiating (stem) cells. In this study, we show that this protein also upregulates the immune checkpoint molecule PD-L1 under conditions of stress in breast and glioblastoma cancer cells, revealing a previously unknown connection between stemness programs, stress responses, and immune checkpoint control. We also identified a significant correlation between G3BP2 and PD-L1 co-expression in tumor tissues from cancer patients. To assess the targetability of G3BP2, we employed a small molecule (C108) that binds G3BP2 and interferes with the stress response. Tumors treated with C108 had increased CD8 T-cell proliferation and infiltration. Moreover, treatment of breast tumor-bearing mice with C108 resulted in a significant survival benefit and long-term cures. Cancer cells treated with C108 or cancer cells with genetically repressed G3BP2 had decreased PD-L1 expression due to enhanced mRNA degradation. Our study provides a compelling mechanism linking stress granule formation and immune checkpoint program of cancer, suggesting this link may provide new opportunities for improving anticancer immunotherapy.

Effects of endogenous proteins on the hydrolysis of gelatinized starch and their mechanism of inhibition

This study investigated the effect of endogenous proteins on the α-amylase-catalyzed hydrolysis of gelatinized corn starch, to analyze the influence of starch granule-associated protein (SGAP) and storage protein on the hydrolytic reaction and the structure of the hydrolysate molecules. The effect of chemicals that disrupted starch/protein binding interactions on the rheology of gelatinized starch revealed the mechanism of starch hydrolysis inhibition. Native corn starch had a lower degree of hydrolysis and a higher proportion of incompletely hydrolyzed long chains than corn starch with the SGAP removed. Storage protein also inhibited starch hydrolysis, but less strongly than SGAP. Rheological analysis showed that SGAP interacted strongly with starch chains by hydrogen bonding, hydrophobic and electrostatic interactions, whereas storage protein only interacted strongly by hydrogen bonding. The stronger binding interactions between SGAP and starch appeared to be the main reason for its stronger inhibition of starch hydrolysis than that of storage protein.

Removal of starch granule-associated proteins alters the physicochemical properties of diverse small granule starches

The properties of four small-granule (oat, quinoa, amaranth and rice) starches before and after removal of starch granule-associated proteins (SGAPs), particularly surface-proteins and channel-proteins, were investigated and compared with two large-granule (waxy and normal maize) starches. The hypothesis, which was confirmed by results, was that the small-granule starches would be more susceptible to water and heat than the large-granule starches after removal of SGAPs on surfaces and in channels using protease. Surface-proteins of some granules in oat and rice starch presented as a film and channel-proteins were observed in oat, quinoa and rice starches. Compared to large-granule starches, removing SGAPs led to a greater reduction in gelatinization temperature for small granule starches. Meanwhile the reduced extent of rigidity of swollen small-granules during shearing was greater after removing SGAPs, corresponding to the reduced peak, breakdown and final viscosities. Furthermore, the removal of SGAPs for small-granule starch pastes weakened pseudoplasticity and shear-thinning behaviour. Removing SGAPs contributed to amylose leaching, accelerated the recrystallization and rearrangement of starch molecules, and facilitated retrogradation, thereby leading to a stronger gel network.

Novel phasins from the Arctic Pseudomonas sp. B14-6 enhance the production of polyhydroxybutyrate and increase inhibitor tolerance

Phasin (PhaP), one of the polyhydroxyalkanoate granule-associated protein, enhances cell growth and polyhydroxybutyrate (PHB) biosynthesis by regulating the number and size of PHB granules. However, few studies have applied phasins to various PHB production conditions. In this study, we identified novel phasin genes from the genomic data of Arctic soil bacterium Pseudomonas sp. B14-6 and determined the role of phaP1Ps under different PHB production conditions. Transmission electron microscopy and gel permeation chromatography revealed small PHB granules with high-molecular weight, while differential scanning calorimetry showed that the extracted PHB films had similar thermal properties. The phasin protein derived from Pseudomonas sp. B14-6 revealed higher PHB production and exhibited higher tolerance to several lignocellulosic biosugar-based inhibitors than the phasin protein of Ralstonia eutropha H16 in a recombinant Escherichia coli strain. The increased tolerance to propionate, temperature, and other inhibitors was attributed to the introduction of phaP1Ps, which increased PHB production from lignocellulosic hydrolysate (2.39-fold) in the phaP1Ps strain. However, a combination of phasin proteins isolated from two different sources did not increase PHB production. These findings suggest that phasin could serve as a powerful means to increase robustness and PHB production in heterologous strains.

Identification of immune related cells and crucial genes in the peripheral blood of ankylosing spondylitis by integrated bioinformatics analysis.

BackgroundAnkylosing spondylitis (AS) is a progressive rheumatic disease and studies reveal that the immune system is critical for the pathogenesis of AS. In the present study, various bioinformatics analysis methods were comprehensively applied, designed to identify potential key genes and inflammation states of AS.MethodsThe transcriptome profiles of GSE25101 and GSE73754 obtained from the Gene Expression Omnibus (GEO) database were merged for subsequent analyses. The differentially expressed genes (DEGs) were identified using the Bioconductor package Limma and threshold values. Functional enrichment and pathway enrichment analyses were performed using the clusterProfiler package and Gene Set Enrichment Analysis (GSEA). Next, protein–protein interaction (PPI) network of the identified DEGs was constructed by the online database, the Search Tool for the Retrieval of Interacting Genes (STRING), visualization and analysis were performed through Cytoscape software. Subsequently, we applied CIBERSORT algorithm to identify subpopulation proportions of immune cells in peripheral blood samples. Finally, we validated the hub genes with the GSE18781 dataset. Samples were collected from patients to validate gene and protein expression using qRT-PCR and ELISA.ResultsA total of 334 DEGs were identified, including 182 upregulated and 152 downregulated DEGs, between AS patients and normal human controls, which were primarily involved in immune response, autophagy, and natural killer cell-mediated cytotoxicity. The most prominent module and candidate biomarkers were identified from the PPI network. Biomarkers were selected for validation and their expressions were significantly decreased in peripheral blood samples which was consistent with transcriptome sequencing results. Nine genes with AUC > 0.70 were considered to be AS hub genes for ROC curve analysis, including GZMA, GZMK, PRF1, GNLY, NKG7, KLRB1, KLRD1, IL2RB and CD247. Furthermore, CIBERSORT results suggest that AS contained a higher proportion of CD8+ T cells, naive CD4+ T cells, neutrophils, and lower levels of gamma delta T cells compared with the normal controls.ConclusionIn this study, we identified DEGs combined with their closely related biological functions and propose that granule-associated proteins and immune infiltration maybe involved in the progression of ankylosing spondylitis. These validated hub genes may provide new perspectives for understanding the molecular mechanisms of ankylosing spondylitis.

Three-dimensional label-free visualization and quantification of polyhydroxyalkanoates in individual bacterial cell in its native state

Polyhydroxyalkanoates (PHAs) are biodegradable polyesters that are intracellularly accumulated as distinct insoluble granules by various microorganisms. PHAs have attracted much attention as sustainable substitutes for petroleum-based plastics. However, the formation of PHA granules and their characteristics, such as localization, volume, weight, and density of granules, in an individual live bacterial cell are not well understood. Here, we report the results of three-dimensional (3D) quantitative label-free analysis of PHA granules in individual live bacterial cells through measuring the refractive index distributions by optical diffraction tomography (ODT). The formation and growth of PHA granules in the cells of Cupriavidus necator, the best-studied native PHA producer, and recombinant Escherichia coli harboring C. necator poly(3-hydroxybutyrate) (PHB) biosynthesis pathway are comparatively examined. Through the statistical ODT analyses of the bacterial cells, the distinctive characteristics for density and localization of PHB granules in vivo could be observed. The PHB granules in recombinant E. coli show higher density and localization polarity compared with those of C. necator, indicating that polymer chains are more densely packed and granules tend to be located at the cell poles, respectively. The cells were investigated in more detail through real-time 3D analyses, showing how differently PHA granules are processed in relation to the cell division process in native and nonnative PHA-producing strains. We also show that PHA granule-associated protein PhaM of C. necator plays a key role in making these differences between C. necator and recombinant E. coli strains. This study provides spatiotemporal insights into PHA accumulation inside the native and recombinant bacterial cells.

Surface microstructure of rice starch is altered by removal of granule-associated proteins

The morphological characteristics and surface structure of waxy, low-amylose and high-amylose rice starches before and after extraction of granule-associated proteins (SGAPs), especially surface-proteins and channel-proteins, were systematically investigated. Removal of SGAPs on surfaces and in channels of granules without significant change in morphological structure was evidenced by microscope images of granules and confocal laser scanning microscope images of fluorescamine-stained and 8-amino-1,3,6-pyrenetrisulfonic acid-stained granules. For nano-scale surface features, atomic force microscopy showed that a more roughened surface occurred on granules after removal of SGAPs, meanwhile, specific surface area, total pore volume, surface and average diameter of pores increased in granules. The results gave direct evidence that extraction of SGAPs induced larger surface area and increased size of channels (pores) in starch granules. Once SGAPs have been removed, dextran-probes could quickly diffuse into channels and cavities, thus probe-diffusion of the granular matrix was accelerated. Although waxy rice starch had the lowest SGAPs content, the change of dextran diffusion into its granule was the greatest of three tested rice starches when SGAPs were removed. The surface texture and porosity of the starch granules before and after removal of SGAPs were significantly correlated to the pasting properties of starch, providing a new insight into the SGAPs-surface microstructure-function relationship of starch granules.

A Critical Role for the IKKβ/Lipid Rafts Axis in Regulating Platelet Exocytosis, Via the Spatial Regulation of SNARE Proteins

Lipid rafts, which are membrane microdomains that are rich in cholesterol and glycosphingolipids, have been implicated in the regulation of intracellular protein trafficking. Furthermore, we have previously shown that SNAREs mediate granule-plasma membrane fusion during exocytosis in platelets, and that IKKβ regulates SNAREs and granule exocytosis. Based on these considerations, we sought to investigate the regulatory relationship between IKKβ and lipid rafts, by employing the PF4-Cre/IKKβflox/flox platelets. We demonstrate that lipid rafts play a critical role in mediating IKKβ-/--dependent regulation of SNARE proteins and SNARE complexes. Specifically, we found that methyl β-cyclodextrin (MβCD, which disrupts lipid rafts) is able to inhibit platelet aggregation, dense and alpha granule secretion, PS exposure and αIIbβ3 integrin activation. We also observed that MβCD is able to inhibit platelet spreading, clot retraction, as well as 7S complex formation. In terms of co-localization, the SNARE protein SNAP-23 was found to associate with a lipid raft marker and was present in detergent-insoluble lipid raft microdomains in control platelets, but not in the IKKβ-/- platelets. Furthermore, in resting platelets, plasma membrane SNARE proteins such as syntaxin-11 and SNAP-23 or the granule-associated SNARE vesicle-associated membrane protein VAMP-8 were neither present in the microdomains of control platelets, nor in those from IKKβ-/- mice. Interestingly however, agonist/thrombin stimulation led to the relocation of syntaxin-11 and VAMP-8 into the lipid rafts, and as part of raft-associated phospho-SNAP-23/syntaxin-11/VAMP-8 complexes in control platelets. This seems to be IKKβ dependent, as these associations did not take place in the IKKβ-/- platelets. Taken together, these data reveal a critical role for the IKKβ/lipid rafts axis in regulating platelet exocytosis, via the spatial regulation of SNARE proteins. DisclosuresNo relevant conflicts of interest to declare.

Effects of drying temperature on the properties of starch in pasta

The effects of manufacturing process and drying temperature on the properties of starch in pasta were investigated. Starch was isolated from durum wheat semolina, raw pasta, and pasta dried under low temperature, high temperature, and very high temperature. The following measurements were carried out on the extracted starch samples: protein content, ash, amylose content, resistant starch, starch damage, swelling power, amount of solubilized starch, X-ray diffraction spectral analysis, gelatinization temperature, granule size distribution, compression behavior of the gels, and the amount of starch granule-associated protein. The starch isolated from each sample had different physicochemical properties. Starch isolated from raw pasta formed a soft gel and had a low amount of starch granule surface-bound protein compared to starch isolated from semolina. In contrast, increasing the drying temperature of pasta increased the hardness and brittleness of gels formed by the extracted starch, and increased the amount of starch granule surface-bound protein. These results suggest that the amount of starch granule surface-bound protein may be an indicator of starch-protein interaction, which affects the physical properties of extracted starch gels.

Impact of starch granule-associated channel protein on characteristic of and λ-carrageenan entrapment within wheat starch granules

This study investigated the physicochemical characteristics of protease-treated wheat starch (PT-WST) to understand the role of starch granule-associated proteins (SGAPs) and the potential capability of PT-WST to provide a nutrient delivery system (NDS). Protease treatment was conducted at 4 °C and 37 °C (PT04 and PT37), respectively. A model delivery system was assessed with PT37 granules infiltrated by λ-carrageenan (λC) under variations of molecular size (λC hydrolysates produced from 0, 2.5, 100, and 500 mM HCl solution), agitation time, and temperature. Protein-specific (3-(4-carboxybenzyl)quioline-2-carboxaldehyde) or non-reactive (methanolic merbromin) fluorescent dye staining revealed that removal of SGAPs on surfaces and channels were more effective for PT37 than for PT04. Consistent amylose content, swelling, and gelatinization temperature before and after protease treatment suggested minimal impact on the starch structure. PT37 presented higher solubility and pasting viscosity than PT04. This resulted from excessive SGAP removal, which enhanced entrapment capacity. λC molecular size and agitation temperature showed a negative correlation with the content of λC entrapped within PT37, and this content depended on the interplay between the agitation time and λC molecular size. As λC molecular size decreased, the λC distribution became uniform throughout the granules, which confirmed the potential of PT-WST as a carrier for NDS.

Recent technological and strategical developments in the biomanufacturing of 1,3-propanediol from glycerol

In the pursuit of sustainable development, there is a rapid increase in the generation of biodiesel, which eventually results in the synthesis of glycerol as a major by-product. The surplus amount of glycerol from biodiesel industries is one of the rising environmental concerns. Alternatively, glycerol can serve as an attractive raw material for synthesizing value-added products that offer simultaneous management and valorization of this waste. 1,3-propanediol is a vital value-added material having extensive applications in the industrial domain. Microbial biosynthesis of 1,3-propanediol from glycerol has proven to be an economic and eco-friendly approach. However, biosynthesis of this bio-product is often affected by some environmental stresses, such as glycerol and 1,3-propanediol concentrations, accumulation of 3-hydroxypropionaldehyde, and by-product synthesis. To overcome the influences of these factors as well as to improve 1,3-propanediol yield, efforts have been devoted over the past years, which mostly included the application of metabolic engineering strategies to transform microbial strains into robust cell factories. This review focuses on the environmental stresses restricting 1,3-propanediol biosynthesis, and how these restrictions could be overcome using the metabolic engineering strategies. In this context, some technological and strategical aspects, including rechanneling metabolic flux in co-producing strains, rebalancing of co-factors and gene expression, co-culture and co-substrate fermentation, and application of untranslated region engineering. It also discusses the role of key enzymes involved in 1,3-propanediol biosynthesis and the influence of polyhydroxyalkanoate granule-associated protein on the tolerance to high concentration of this bio-product.

ALS-Linked Mutant SOD1 Associates with TIA-1 and Alters Stress Granule Dynamics.

Amyotrophic lateral sclerosis (ALS) is a degenerative disorder caused by motor neuron loss. T-cell intracellular antigen-1 (TIA-1), a cytotoxic T lymphocyte granule-associated RNA binding protein, is a key component of stress granules. However, it remains uncertain whether ALS-causing superoxide dismutase-1 (SOD1) toxicity alters the dynamics of stress granules. Thus, through mouse and cell line models, and human cells and tissues, we showed the subcellular location of TIA-1 and its recruitment by stress granules following mutant SOD1-related stimuli. An overexpression of MTSOD1 resulted in increased TIA-1-positive cytoplasmic inclusions in the spinal cord tissue of SOD1G93A transgenic mouse and the SOD1G86S familial ALS patient. Moreover, we demonstrated the stages of ALS-like disease-dependent increase in TIA-1 in the spinal cord of transgenic mice. A similar increase of TIA-1 was found in the spinal cord of the SOD1G86S patient and induced pluripotent stem cell-derived neural stem cells from the SOD1G17S patient. By using immunoprecipitation assays in wild type (WT) human SOD1 (hSOD1) or mutant (MT) hSOD1-transfected motor neuronal cell lines and SOD1G93A transgenic mouse model, we observed that MTSOD1 interacts with TIA-1. In WT or MT hSOD1-transfected HEK293 and NSC-34 cells, the formation of TIA-1-positive stress granules was delayed in MTSOD1 by sodium arsenite treatment. These findings suggest that MTSOD1 could affect the dynamics of stress granules through the abnormal MTSOD1-TIA-1 interaction. Consequently, the resulting pathological TIA-1 may be involved in RNA metabolism found in ALS.

Impact of starch granule-associated surface and channel proteins on physicochemical properties of corn and rice starches

The objective of this study was to investigate the impact of removing starch granule-associated proteins (SGAPs), especially starch granule-associated surface and channel proteins, on the overall characteristics of corn and rice starches. Protease treatment predominantly removed SGAPs on surfaces and in channels of the starches without significant damage, as evidenced by confocal laser scanning microscopy coupled with protein-specific and non-reactive fluorescent dye staining. Compared to untreated starches, protease-treated (PT) starches showed higher solubility and lower swelling power. However, there were no changes in their gelatinization and melting temperatures, despite their higher relative crystallinity. The stability of swollen starch granules during shearing was reduced following SGAP removal, reducing their peak, final and setback viscosities of the paste. Taken together these results indicate that SGAP removal predominantly affects the rheological properties of starch. Furthermore, the lower setback in PT-starch pastes suggests short-term retrogradation may be retarded by protease treatment of starch.

Removal of starch granule-associated proteins affects amyloglucosidase hydrolysis of rice starch granules

Starch granule-associated proteins (SGAPs) include granule-surface proteins (SGSPs) and granule-channel proteins (SGCPs). To investigate impacts of SGAPs on amyloglucosidase (AMG) hydrolysis, waxy and non-waxy rice starches had their SGCPs or SGAPs removed. Removal of SGAPs or SGCPs did not affect morphology and amylopectin chain distribution but decreased relative crystallinity. Removal of SGAPs increased the digestion rate, AMG binding ability and pore diameter of hydrolyzed granules, and accelerated changes in relative crystallinity and destruction of crystalline region on hydrolysis. However, after removing SGCPs, AMG only bound to surface and attacked of the fingerprint of protein bodies on granules, with decreased hydrolysis rate. The degree of change in hydrolysis rate was not determined by SGCPs content of rice starch. These results implied that SGCPs had a more dominant role in AMG hydrolysis of rice starch than did SGSPs. This study provides novel information about the role of SGAPs in AMG hydrolysis mechanisms.