Search

In the U.S., 30–40% of all food produced ends up as waste. By taking advantage of biological processes this waste nutrients can be converted into value-added 2,3-butanediol (2,3-BDO). However, food waste contains live bacteria and fungi which must first be inactivated, a process typically done with autoclaving under high pressure steam. If contaminants are not removed they can out-compete added industrial microbes for nutrients, resulting in lower yields of bio-products. Unfortunately, sterilization methods such as autoclaving greatly increase production time, and increase energy consumption. This creates a demand for novel microorganisms that can out-compete food waste microbiota by natural means without the need for sterilization of food waste. For this study, several Bacillus licheniformis isolates from Yellowstone National Park were investigated for their potential use in the 2,3-BDO industry. B. licheniformis species are a strain of industrial importance as of late, due to the wide metabolic functions and sustainable growth conditions above 45 °C. Whole genomic sequencing data and biochemical classification of B. licheniformis YNP1-TSU, B. licheniformis YNP2-TSU, and B. licheniformis YNP3-TSU were assessed and revealed a wide assortment of anti-microbial peptides and antibodies as well as catabolic genes for assimilation of glucose, fructose, galactose, xylose, arabinose, mannose, starch and sucrose, and linoleic acid. All three B. licheniformis strains had an optimal growth temperature of 50 °C, and were identified as high 2,3-BDO producers. Each strain YNP1-TSU, YNP2-TSU, and YNP3-TSU yielded 0.44 g/g, 0.45 g/g, and 0.43 g/g from mixed sugars found in non-sterilized food waste, respectively.

Lignocellulosic biomass is the earth’s most abundant renewable feedstock alternative that comprises of cellulose, hemi-cellulose and lignin. The synergistic action of cellulolytic/xylanolytic enzymes produced by lignocellulolytic micro-organisms such as bacteria, algae and fungi are capable of robust cellulosic biomass deconstruction. Most of the microorganisms dwelling in extreme environmental habitats such as rumen environment, hot/cold springs, deep ocean trenches, acidic/alkaline pH environment have been considered as an attractive producers of hemi/cellulolytic lignocellulolytic and other biotechnological enzymes with enhanced bio-chemical properties essential for industrial bioconversion processes. However, the potential microbial sources of cellulolytic enzymes and the underlying mechanism to achieve this is not fully elucidated. In this review article, first we detail the composition of lignocellulosic biomass. Next, we describe the structure and functions of divergent hydrolytic enzymes (cellulolytic and xylanolytic enzymes) involved in cellulosic biomass degradation. Third, we analyze, outline and unveil the prospective source of microbes encoding exceptionally diverse set of biotechnologically relevant cellulolytic enzymes which are critical to answer the specific ecological question of by whom, where and how cellulosic biomass is degraded in the environment. Finally, this review article features the snapshot about the future developments and perspectives on microbial enzymes, high-throughput techniques and molecular tools that could be exploited to derive those enzymes from the potential sources.

Microbes are ubiquitously distributed in nature and are a critical part of the holobiont fitness. They are perceived as the most potential biochemical reservoir of inordinately diverse and multi-functional enzymes. The robust nature of the microbial enzymes with thermostability, pH stability and multi-functionality make them potential candidates for the efficient biotechnological processes under diverse physio-chemical conditions. The need for sustainable solutions to various environmental challenges has further surged the demand for industrial enzymes. Fueled by the recent advent of recombinant DNA technology, genetic engineering, and high-throughput sequencing and omics techniques, numerous microbial enzymes have been developed and further exploited for various industrial and therapeutic applications. Most of the hydrolytic enzymes (protease being the dominant hydrolytic enzyme) have broad range of industrial uses such as food and feed processing, polymer synthesis, production of pharmaceuticals, manufactures of detergents, paper and textiles, and bio-fuel refinery. In this review article, after a short overview of microbial enzymes, an approach has been made to highlight and discuss their potential relevance in biotechnological applications and industrial bio-processes, significant biochemical characteristics of the microbial enzymes, and various tools that are revitalizing the novel enzymes discovery.

Background Bacillus cereus is a bacterial species which grows efficiently on a wide range of carbon sources and accumulates biopolymer poly-hydroxybutyrate (PHB) up to 80% cell dry weight. PHB is an aliphatic polymer produced and stored intracellularly as a reservoir of carbon and energy, its mobilization is a key biological process for sporulation in Bacillus spp. Previously, B. cereus tsu1 was isolated and cultured on rapeseed cake substrate (RCS), with maximum of PHB accumulation reached within 12 h, and depleted after 48 h. Fore-spore and spore structure were observed after 24 h culture. Results Quantitative proteomic analysis of B. cereus tsu1 identified 2952 quantifiable proteins, and 244 significantly changed proteins (SCPs) in the 24 h:12 h pair of samples, and 325 SCPs in the 48 h:12 h pair of samples. Based on gene ontology classification analysis, biological processes enriched only in the 24 h:12 h SCPs include purine nucleotide metabolism, protein folding, metal ion homeostasis, response to stress, carboxylic acid catabolism, and cellular amino acid catabolism. The 48 h:12 h SCPs were enriched into processes including carbohydrate metabolism, protein metabolism, oxidative phosphorylation, and formation of translation ternary structure. A key enzyme for PHB metabolism, poly(R)-hydroxyalkanoic acid synthase (PhaC, KGT44865) accumulated significantly higher in 12 h-culture. Sporulation related proteins SigF and SpoEII were significantly higher in 24 h-samples. Enzymes for nitrate respiration and fermentation accumulated to the highest abundance level in 48 h-culture. Conclusions Changes in proteome of B. cereus tsu1 during PHB intracellular mobilization were characterized in this study. The key enzyme PhaC for PHB synthesis increased significantly after 12 h-culture which supports the highest PHB accumulation at this time point. The protein abundance level of SpoIIE and SigF also increased, correlating with sporulation in 24 h-culture. Enzymes for nitrate respiration and fermentation were significantly induced in 48 h-culture which indicates the depletion of oxygen at this stage and carbon flow towards fermentative growth. Results from this study provide insights into proteome profile changes during PHB accumulation and reuse, which can be applied to achieve a higher PHB yield and to improve bacterial growth performance and stress resistance.

Cotton (Gossypium spp.) is the world’s third most important oilseed crop as well as an economic source of protein. However, the presence of (-)- gossypol limits the nutritional value of cottonseeds. To genetically profile relevant traits, seeds of the two cotton parental lines TM-1 (G. hirstum) and 3-79 (G. barbadense) along with their 17 chromosomal substitution (CS) progenies were quantified for (-)- and (+)- enantiomer levels. The TM-1 parent was found to contain a higher ratio of (+)-:(-)- gossypol enantiomers (13.34:8.52 µg/mg) compared to that of 3-79 levels (6.69:7.41 µg/mg) in the seeds. Among progeny CS-B (containing substituted chromosomes from G. barbadense in G. hirsutum background) lines, only CS-B17 was found to contain significantly lower (7.65 µg/mg) level of (+)-enantiomer in its seeds, and also had the lowest (5.09 µg/mg) amount of (-)- gossypol among all lines. A singular protocol was then devised for RNA yields from polysaccharide-rich cottonseeds of CS-B17 and its parents. Transcription derived fragments (TDFs) analyses of these three lines were conducted on seeds harvested at 28, 35 and 42 days post anthesis (DPA). 64 TDFs profiles were analyzed using differential expression analyses, and similarities were revealed between 3-79 and CS-B17 along with unique expression trends for CS-B17. Such expression analyses, which are based on single chromosome differences with TM-1 parent while reflecting seed nutritional merit of 3-79 parent, may facilitate cotton breeding endeavors. Keywords: Chromosome substituted lines; Cotton; High performance liquid chromatography; Nutrition traits; RNA extraction; Transcription derived fragments http://dx.doi.org/10.19045/bspab.2019.80129

Optimization of cellulase activity is vital for synthesizing the end-products of second generation biofuel production. The slightest change in fermentation parameters can reduce the secretion of necessary enzymes to degrade cellulosic biomass. Determining the ecological effects of certain key media components is essential to understand how bacterial species will respond in a fluid environment. For our experiment a cellulosic media was designed to enhance the industrially important thermophile, Bacillus licheniformis YNP5-TSU. After several attempts to simplify the carboxymethylcellulose (CMC) media composition, impaired biofilm maturation and cellulase activity was noticed. This negative artifact occurred only when magnesium sulphate was removed from media. To analyze the shift in gene expression caused by magnesium stress, biofilm associated proteins were extracted from both control (4.0 mM MgSO4) and magnesium depleted (0.0 mM MgSO4) media at 24 hr and 48 hr incubation periods. These proteins were quantified through isobaric labeling and raw data generated from nanoLC-MS/MS identified over 2,000 proteins from the Bacillus licheniformis YNP5-TSU proteome (NCBI accession number MEDD00000000). After statistical normalization and false discovery rate were calculated, a total of 161 proteins from magnesium depleted media and 238 proteins from control media were deemed statistically relevant. A closer look through STRING interconnected webs, data mining, and NCBI annotations revealed several up/down regulated proteins that had linkage to biofilm formation and cellulase secretion. In this study we are able to provide significant evidence that; (1) biofilm maturation and cellulase production are highly correlated and (2), their optimization is dependent on the expression of several key proteins.

The complete affiliations of three authors and one of the co-author’s name, “Naveen Kumar Arora” was published incorrectly in the original publication of the article. The correct version of affiliations and the co-author’s name has been corrected in this correction.

This paper reports a laser capture microdissection-tandem mass tag-quantitative proteomics analysis of Al-sensitive cells in root tips. Cherry tomato (Solanum lycopersicum var. cerasiforme ‘LA2710’) seedlings were treated under 15 μM Al3+ activity for 13 d. Root-tip longitudinal fresh frozen tissue sections of 10 μm thickness were prepared. The Al-sensitive root zone and cells were determined using histochemical analysis of root-tips and micro-sections. A procedure for collecting the Al-sensitive cells using laser capture microdissection-protein extraction-tandem mass tag-proteomics analysis was developed. Proteomics analysis of 18 μg protein/sample with three biological replicates per treatment condition identified 3879 quantifiable proteins each associated with two or more unique peptides. Quantified proteins constituted a broad range of Kyoto Encyclopedia of Genes and Genomes pathways when searched in the annotated tomato genome. Differentially expressed proteins between the Al-treated and non-Al treated control conditions were identified, including 128 Al-up-regulated and 32 Al-down-regulated proteins. Analysis of functional pathways and protein-protein interaction networks showed that the Al-down-regulated proteins are involved in transcription and translation, and the Al-up-regulated proteins are associated with antioxidant and detoxification and protein quality control processes. The proteomics data are available via ProteomeXchange with identifier PXD010459 under project title ‘LCM-quantitative proteomics analysis of Al-sensitive tomato root cells’. Significance This paper presents an efficient laser capture microdissection-tandem mass tag-quantitative proteomics analysis platform for the analysis of Al sensitive root cells. The analytical procedure has a broad application for proteomics analysis of spatially separated cells from complex tissues. This study has provided a comprehensive proteomics dataset expressed in the epidermal and outer-cortical cells at root-tip transition zone of Al-treated tomato seedlings. The proteomes from the Al-sensitive root cells are valuable resources for understanding and improving Al tolerance in plants.

Glyoxalase I (Gly I) is the first enzyme in the glutathionine-dependent glyoxalase pathway for detoxification of methylglyoxal (MG) under stress conditions. Transgenic tomato ‘Money Maker’ plants overexpressing tomato SlGlyI gene (tomato unigene accession SGN-U582631/Solyc09g082120.3.1) were generated and homozygous lines were obtained after four generations of self-pollination. In this study, SlGlyI-overepxressing line (GlyI), wild type (WT, negative control) and plants transformed with empty vector (ECtr, positive control), were subjected to Al-treatment by growing in Magnavaca’s nutrient solution (pH 4.5) supplemented with 20 µM Al3+ ion activity. After 30 days of treatments, the fresh and dry weight of shoots and roots of plants from Al-treated conditions decreased significantly compared to the non-treated conditions for all the three lines. When compared across the three lines, root fresh and dry weight of GlyI was significant higher than WT and ECtr, whereas there was no difference in shoot tissues. The basal 5 mm root-tips of GlyI plants expressed a significantly higher level of glyoxalase activity under both non-Al-treated and Al-treated conditions compared to the two control lines. Under Al-treated condition, there was a significant increase in MG content in ECtr and WT lines, but not in GlyI line. Quantitative proteomics analysis using tandem mass tags mass spectrometry identified 4080 quantifiable proteins and 201 Al-induced differentially expressed proteins (DEPs) in root-tip tissues from GlyI, and 4273 proteins and 230 DEPs from ECtr. The Al-down-regulated DEPs were classified into molecular pathways of gene transcription, RNA splicing and protein biosynthesis in both GlyI and ECtr lines. The Al-induced DEPs in GlyI associated with tolerance to Al3+ and MG toxicity are involved in callose degradation, cell wall components (xylan acetylation and pectin degradation), oxidative stress (antioxidants) and turnover of Al-damaged epidermal cells, repair of damaged DNA, epigenetics, gene transcription, and protein translation. A protein–protein association network was constructed to aid the selection of proteins in the same pathway but differentially regulated in GlyI or ECtr lines. Proteomics data are available via ProteomeXchange with identifiers PXD009456 under project title ‘25Dec2017_Suping_XSexp2_ITAG3.2’ for SlGlyI-overexpressing tomato plants and PXD009848 under project title ‘25Dec2017_Suping_XSexp3_ITAG3.2’ for positive control ECtr line transformed with empty vector.

Pollen development is highly susceptible to heat stress (HS) and the production of inviable pollen causes reduction in seed- and fruit-set in plants. This study was carried out to identify HS-induced pollen proteins and the associated biological processes in tomato (Solanum lycopersicum). Tomato ‘Micro-Tom’ plants were incubated under 32°C//22°C (day/night, 12/12 h) for two weeks for heat treatment, and the non-treated control plants were incubated for the same time period at 25°C /22°C. Flower buds of 5 mm in length were confirmed to contain the heat sensitive uninucleate microspores. Pollen cells were harvested using laser capture microdissection (LCM) and protein was extracted using a one-step method under high pressure and vacuum. Approximately 60,000 LCMharvested microspore cells yielded about 18-20 μg proteins. The tandem mass tags (TMT) proteomics analysis identified a total of 6018 proteins, 4784 proteins were quantified, 37 proteins were identified as HS up-regulated significantly changed proteins (SCPs), and 83 proteins as HS down (dn)-regulated SCPs. Further analysis using the plant MetGenMap system showed that the HS up-regulated SCPs were enriched in the heat acclimation, pollen wall formation, protein folding/refolding gene ontology (GO) biological processes, and the HS dn-regulated SCPs were placed in the carbohydrate catabolism and de-novo protein biosynthesis GO terms. Biological processes such as mitosis, resistance to oxidative stresses, and carbohydrate and lipid metabolic processes contain both the HS up-, and dn-regulated SCPs. These results indicate that the LCM-TMT proteomics workflow is highly efficient in the identification of HS-induced pollen proteomes. These HS induced SCPs will be used for exploring heat tolerance of tomato pollens. The proteomics data are available via ProteomeXchange with identifier PXD010218.

In this paper, we report on aluminum (Al)-induced root proteomic changes in switchgrass. After growth in a hydroponic culture system supplemented with 400 μM of Al, plants began to show signs of physiological stress such as a reduction in photosynthetic rate. At this time, the basal 2-cm long root tips were harvested and divided into two segments, each of 1-cm in length, for protein extraction. Al-induced changes in proteomes were identified using tandem mass tags mass spectrometry (TMT-MS)-based quantitative proteomics analysis. A total of 216 proteins (approximately 3.6% of total proteins) showed significant differences between non-Al treated control and treated groups with significant fold change (twice the standard deviation; FDR adjusted p-value < 0.05). The apical root tip tissues expressed more dramatic proteome changes (164 significantly changed proteins; 3.9% of total proteins quantified) compared to the elongation/maturation zones (52 significantly changed proteins, 1.1% of total proteins quantified). Significantly changed proteins from the apical 1-cm root apex tissues were clustered into 25 biological pathways; proteins involved in the cell cycle (rotamase FKBP 1 isoforms, and CDC48 protein) were all at a reduced abundance level compared to the non-treated control group. In the root elongation/maturation zone tissues, the identified proteins were placed into 18 pathways, among which proteins involved in secondary metabolism (lignin biosynthesis) were identified. Several STRING protein interaction networks were developed for these Al-induced significantly changed proteins. This study has identified a large number of Al-responsive proteins, including transcription factors, which will be used for exploring new Al tolerance genes and mechanisms. Data are available via ProteomeXchange with identifiers PXD008882 and PXD009125.

This study aimed to identify the aluminum (Al)-induced proteomes in tomato (Solanum lycopersicum, "Micro-Tom") after long-term exposure to the stress factor. Plants were treated in Magnavaca's solution (pH 4.5) supplemented with 7.5 μM Al(3+) ion activity over a 4 month period beginning at the emergence of flower buds and ending when the lower mature leaves started to turn yellow. Proteomes were identified using a 8-plex isobaric tags for relative and absolute quantification (iTRAQ) labeling strategy followed by a two-dimensional (high- and low-pH) chromatographic separation and final generation of tandem mass spectrometry (MS/MS) spectra of tryptic peptides on an LTQ-Orbitrap Elite mass spectrometer. Principal component analysis revealed that the Al-treatment had induced systemic alterations in the proteomes from roots and leaves but not seed tissues. The significantly changed root proteins were shown to have putative functions in Al(3+) ion uptake and transportation, root development, and a multitude of other cellular processes. Changes in the leaf proteome indicate that the light reaction centers of photosynthetic machinery are the primary targets of Al-induced stress. Embryo and seed-coat tissues derived from Al-treated plants were enriched with stress proteins. The biological processes involving these Al-induced proteins concur with the physiological and morphological changes, such as the disturbance of mineral homeostasis (higher contents of Al, P, and Fe and reduced contents of S, Zn, and Mn in Al-treated compared to nontreated plants) in roots and smaller sizes of roots and the whole plants. More importantly, the identified significant proteins might represent a molecular mechanism for plants to develop toward establishing the Al tolerance and adaptation mechanism over a long period of stress treatment.

Strawberry production and consumption is experiencing significant growth. Production in the USA has doubled since the 1990s. As the domestic demand increases, there is potential to expand production beyond California and Florida into other regions of the country. In the 1950s, strawberries accounted for 25.3 % of the total marketable value of all fruits and vegetables produced in Tennessee, before declining in the 1970s. Acreage in the last 2 decades has been on a slow upward trend. Several counties throughout Tennessee are well suited for the growing of high-quality and high-value strawberries. Efforts are being made to promote production and consumption of organic and naturally grown local strawberries in Tennessee. Strawberry production in parts of China that have a similar climate has also seen considerable growth. Hebei, Shandong, Anhui, and Liaoning are among the largest strawberry producing provinces in the country. In 2014, over 50,000 acres of strawberries were planted in Anhui Province, accounting for over 15 % of all strawberries grown in China. This chapter discusses the production systems, cultural practices including nutrient management, the selection of cover crops, integrated pest management, and weed and disease control in strawberry production in Tennessee and China. Harvesting and postharvesting protocols, marketing channels, safety concerns, current constraints, and future potentials of organic strawberry production are also discussed.

Single-cell-type proteomics provides the capability to revealing the genomic and proteomics information at cell-level resolution. However, the methodology for this type of research has not been well-developed. This paper reports developing a workflow of laser capture microdissection (LCM) followed by gel-liquid chromatography-tandem mass spectrometry (GeLC-MS/MS)-based proteomics analysis for the identification of proteomes contained in individual cell layers of tomato roots. Thin-sections (~10-μm thick, 10 sections per root tip) were prepared for root tips of tomato germinating seedlings. Epidermal and cortical cells (5000-7000 cells per tissue type) were isolated under a LCM microscope. Proteins were isolated and then separated by SDS-polyacrylamide gel electrophoresis followed by in-gel-tryptic digestion. The MS and MS/MS spectra generated using nanoLC-MS/MS analysis of the tryptic peptides were searched against ITAG2.4 tomato protein database to identify proteins contained in each single-cell-type sample. Based on the biological functions, proteins with proven functions in root hair development were identified in epidermal cells but not in the cortical cells. Several of these proteins were found in Al-treated roots only. The results demonstrated that the cell-type-specific proteome is relevant for tissue-specific functions in tomato roots. Increasing the coverage of proteomes and reducing the inevitable cross-contamination from adjacent cell layers, in both vertical and cross directions when cells are isolated from slides prepared using intact root tips, are the major challenges using the technology in proteomics analysis of plant roots.