Amino Acid Composition Analysis Research Articles

A New Method of Gelatin Modified Collagen and Viscoelastic Study of Gelatin-Collagen Composite Hydrogel

Pure collagen materials are expensive with poor mechanical properties, which need modifications in most cases. As the degradation product of collagen, gelatin is cheap, degradable and biocompatible, but few literatures have reported the research about gelatin-collagen composite materials. This is because gelatin and collagen have different soluble temperatures—gelatin is soluble in hot water (≥30 °C) and swells in cold water. However, a low temperature (2–10 °C) is required to prepare and store collagen solution, and neutral collagen solution denatures quickly above the room temperature. In this study, gelatin was ground into powders and swelled in neutral bovine tendon pepsin-soluble collagen solution (BPSC) to form a homogeneous gelatin-collagen mixture, in light of the swelling characteristics of gelatin in cold water. The assembly properties and gel properties of this composite material were further studied. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) test results showed that the bovine tendon collagen had typical type-I collagen structural characterizations with two α chains of about 100 kDa and one β chain of about 200 kDa; while the SDS-PAGE pattern of gelatin displayed bands continuously distributed from 30 to 200 kDa. Amino acid composition analysis test indicated that the content of polar amino acids and the sum of acidic and base amino acids for gelatin were higher than that of BPSC. Studies on gel properties demonstrated that gelatin-collagen mixed solution had collagen-like assembly characteristics and assembly kinetics. The moduli of the assembled gel at 35 °C were equivalent to that of pure bovine tendon collagen system; moreover, the system moduli didn’t change with time with elastic moduli (G′) of about 40 Pa. However, at 25 °C, the moduli of gelatin-collagen composite hydrogel increased with the extension of time, its G′ increased about 18 times within 8 h, and the ratio of elastic modulus to viscous modulus (G″) increased 4.6 times, showing a significant aging effect of structural strength. Meanwhile, the mechanical strength of the composite hydrogel was also regulated by temperature—the gel was highly elastic (G′≈3,000 Pa, G′>>G″) at a low temperature (5 °C); as the temperature rose, the system moduli gradually decreased and the elastic gel transformed into waterlike fluid at 50 °C little by little. What’s more, gelatin-collagen composite hydrogel also had reversible sol-gel performances and self-healing capability similar to the gelatin hydrogel. This novel preparation method for preparing composite materials and the resultant composite hydrogel are expected to be used in the fields of natural food gels, injectable hydrogels, cell scaffolds, drug sustained-release materials and so on, and improve and promote the processing performances, price and large-scale production of collagen-based materials.

Сравнительный анализ аминокислотного состава белка семян подсолнечника селекции ФГБНУ ФНЦ ВНИИМК

Сравнительный анализ аминокислотного состава белка семян подсолнечника селекции ФГБНУ ФНЦ ВНИИМК

Determination of free amino acids in five commercial sweetpotato cultivars by hydrophilic interaction liquid chromatography-mass spectrometry

A sensitive and rapid method was developed for the determination of free amino acid profiles in sweetpotatoes. The method utilized an ultra-high-performance liquid chromatography system with hydrophilic interaction liquid chromatography (HILIC) separation coupled with tandem mass-spectrometry (MS/MS) detection without the need for chemical derivatization. Separation of 36 amino acid standards was achieved on a silica HILIC column in a single MS/MS run of 19 min. This validated method was applied for the analysis of the free amino acid composition of five commercial sweetpotato cultivars (Covington, Beauregard, Hatteras, Murasaki-29 and O’ Henry) from two planting lots grown in experimental fields. Analysis of variance with Tukey’s test was used to determine the differences in individual free amino acid content among the analyzed samples, and principle component analysis (PCA) was implemented to evaluate the variation in amino acid profiles of different sweetpotato cultivars. Asparagine was the most abundant free amino acid and its content varied significantly among the cultivars (p < 0.05). Fifteen additional amino acids also varied among cultivars and contributed to the differentiation of the genotypes by PCA based on their complete amino acid profiles. The efficient, reliable, sensitive method described herein could be used in quantifying amino acids in food matrices similar to sweetpotatoes.

Meat Quality, Amino Acid, and Fatty Acid Composition of Liangshan Pigs at Different Weights

Simple SummaryThe research on the quality of traditional pork can not only provide a reference for the thorough breeding and food development of pigs, but also make a reference for understanding the local history and social culture. The Liangshan pig is a traditional Chinese miniature pig breed. It is mainly raised in the Liangshan Yi area and is closely related to the dietary culture of the local people. The characteristics of, and changes in, the meat quality, amino acid composition and fatty acid composition of Liangshan pigs of different weights were revealed for the first time in this paper. It was found that as the weight of Liangshan pigs increased, the contents of marbling score, intramuscular fat, shear force, Met, Asp, Asn, C18: 0 and C20: 2 increased, and drip loss, Trp and C22: 6 decreased. Taken together, our findings serve as a reference for the development of the local Liangshan pig industry.Indigenous pig breeds are important biological resources and their diversity has been severely damaged. The Liangshan pig is a typical mountain-type local pig breed in southwest China. Here, the meat quality, amino acid, and fatty acid composition of Liangshan pigs were compared at seven stages within the weight range of 50–90 kg. A score for comprehensive factors of meat quality was maintained after rising and kept in a plateau within 74.9–91.5 kg of body weight. The total amount of amino acids in the longissimus dorsi muscle remained stable, and the total fatty acids showed an upward trend. Amino acid composition analysis revealed that as the body weight of Liangshan pigs increased, umami, basic, and acidic amino acid contents decreased, while the essential amino acids (EAA) content and the ratio of basic amino acids to acidic amino acids increased. Fatty acid composition analysis revealed that as body weight increased, the content of polyunsaturated fatty acids (PUFA) exhibited a downward trend, while the content of saturated fatty acids (SFA) exhibited an upward trend. This study is a primary step towards the development and utilization of Liangshan pigs and provides useful information for local pork processing and genetic improvement.

Preparation, physicochemical characterization and identification of two novel mixed ACE-inhibiting peptides from two distinct tea alkali-soluble protein

In this study, two novel mixed ACE-inhibiting peptides, namely Se-TAPep 1 and TAPep 1, were prepared and purified from selenium-enriched and common alkali-soluble proteins using a series of process, and then its physicochemical properties and oligopeptide sequence were analyzed. The results showed that Se-TAPep 1 and TAPep 1 could be successfully separated, but their ACE inhibitory activities were different (IC50 values were 13.39 mg/mL and 23.56 mg/mL, respectively), this may be related to the difference in selenium content. Further analysis of physicochemical properties showed that the protein or peptide fractions contained their characteristic absorption peaks and contained aromatic amino acids, while the apparent morphology was different, the presence of selenium could change the apparent morphology of protein or polypeptide fractions, arranging of selenium-enriched fractions more irregular. The amino acid composition analysis showed that the Pro content of the selenium-enriched fractions was higher than that of the ordinary component, whereas the oligopeptide sequence analysis showed that both LQPSLGFP derived from Se-TAPep 1 and AETGEIKGHY derived from TAPep 1 contained some characteristics of highly active ACE-inhibiting peptides, but the characteristics of LQPSLGFP were more conspicuous, which may be related to its molecular weight and source of mixed peptides. These findings indicated that the selenium-enriched tea mixed peptides had better potential uses than ordinary tea mixed peptides in the preparation of functional food.

Bone-strengthening effects and safety of compound peptides from skin of Chiloscyllium plagiosum and Mustelus griseus.

Fish processing produces a lot of by‐products highly containing large amount of proteins which mainly consist of collagen, implying great potential value for application as nutraceutical ingredients. In present study, two kinds of sharks, Chiloscyllium plagiosum and Mustelus griseus, were used as raw material to gain three kinds of “compound peptides” (CPs) by enzymolysis, FCP (CPs from the flesh of C. plagiosum), SCP (CPs from the skin of C. plagiosum), and SMG (CPs from the skin of M. griseus). According to a series of constituent analysis, the molecule weights of FCP, SCP, and SMG were under 800 Da; amino acids composition analysis of FCP, SCP, and SMG showed that there were high glycine, proline, and hydroxyproline and low cysteine contents in SCP and SMG, which is the characteristic of collagen peptides; their total protein contents were 87.500%, 91.875%, and 95.625%, respectively; and heavy metal contents of CPs were all beneath national standards. After three kinds of CPs were administrated intragastrically to C57BL/6 mice at a total dosage of 15 g/kg, bone‐strengthening effects of SCP and SMG were manifested by osteoblasts activity promotion, bone mineral density (BMD) increase, and marrow adipocyte number decrease, yet nonsignificant effects were shown in FCP group. No index showed toxicity of SCP and SMG in subacute toxicology trial, indicating their safety as functional foods. Herein, industrial application foundation of the skins from these two sharks was explored but more efforts should subsequently be implemented for further exploitation.

A novel thermal hydrolysis process for extraction of keratin from hog hair for commercial applications

We have developed a novel thermal hydrolysis process (THP) to extract and hydrolyze keratin from keratinous animal body wastes without using any chemicals. Our process consists of two heating steps: one is to swell and denature the keratin protein network in the intermediate filaments, while the other is to cleavage the disulfide bonds that connect the tight keratinous fibrils together. Using hog hair as an example, the two-step process achieved a nearly 70% keratin recovery yield, with respect to the original keratin in the hog hair, which is comparable to one of the best recovery yields by conventional chemical processes. The extracted keratin hydrolysate by THP was filtered by the shear wave-induced membrane ultrafiltation for characterization. The molecular weight (MW) analysis using SDS-PAGE and MALDI-TOF mass spectroscopy has demonstrated that our keratin hydrolysis obtained by our two-step THP has a wide range of MW distribution, similar to those already in the hair-care product market. The amino acid composition analysis has shown that our keratin hydrolysate by THP had twice as much essential amino acids as soybean meals on a dry mattter basis. As to the cysteine residue content, we have shown that it can be controlled by adjusting the 2nd heating temperature.

Comparison of the Structural Characteristics of Native Collagen Fibrils Derived from Bovine Tendons using Two Different Methods: Modified Acid-Solubilized and Pepsin-Aided Extraction.

Native collagen fibrils (CF) were successfully extracted from bovine tendons using two different methods: modified acid-solubilized extraction for A-CF and pepsin-aided method for P-CF. The yields of A-CF and P-CF were up to 64.91% (±1.07% SD) and 56.78% (±1.22% SD) (dry weight basis), respectively. The analyses of both amino acid composition and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) confirmed that A-CF and P-CF were type I collagen fibrils. Both A-CF and P-CF retained the intact crystallinity and integrity of type I collagen’s natural structure by FTIR spectra, circular dichroism spectroscopy (CD) and X-ray diffraction detection. The aggregation structures of A-CF and P-CF were displayed by UV–Vis. However, A-CF showed more intact aggregation structure than P-CF. Microstructure and D-periodicities of A-CF and P-CF were observed (SEM and TEM). The diameters of A-CF and P-CF are about 386 and 282 nm, respectively. Although both A-CF and P-CF were theoretically concordant with the Schmitt hypothesis, A-CF was of evener thickness and higher integrity in terms of aggregation structure than P-CF. Modified acid-solubilized method provides a potential non-enzyme alternative to extract native collagen fibrils with uniform thickness and integral aggregation structure.

In vitro antioxidant and anti-aging properties of swim bladder peptides from Atlantic cod (Gadus morhua)

ABSTRACT Two bioactive peptide fractions (SWP-I and SWP-II) of swim bladder peptides from Atlantic cod (Gadus morhua) were obtained through enzymatic hydrolysis and isolated by gel filtration chromatography. The properties of two fractions were characterized by FT-IR, UV scanning spectroscopy, amino acid composition analysis and molecular weight determination. The results showed that the content of aromatic amino acids in SWP-II was higher than that in SWP-I, and the molecular of SWP-I and SWP-II were 4976 Da and 1960 Da, respectively. The radical scavenging activities showed that SWP-I and SWP-II were effective scavengers of DPPH•, HO•, and O2-•, and demonstrated high Fe2+-chelating activity. The scavenging capacities of SWP-II were stronger than those of SWP-I at the same concentration. Cytoprotective experiment suggested that SWP-I and SWP-II possessed ROS-scavenging activity. Pretreatment with these two peptides increased viability rates, suppressed SA-β-gal activities and inhibited apoptosis rates in hydrogen peroxide (H2O2)-induced premature senescent 2BS cells. In conclusion, the enzymatic peptides from swim bladders of Atlantic cod (Gadus morhua) possessed antioxidant activities and could effectively protect premature cell senescence induced by H2O2.

Engraulis encrasicolus larvae from two different environmental spawning areas of the Central Mediterranean Sea: first data on amino acid profiles and biochemical evaluations

Early life stages of marine fish populations may be strongly affected by environmental factors. Changes in the physical environment or the availability of food resources could lead to stress-related physiological responses affecting larval fitness, growth and survival. In the present study, we determined, for the first time, amino acid composition (AAC), lipid, and carbohydrate content, as well as alkaline phosphatase and peroxidase activities in larvae from the European anchovy Engraulis encrasicolus. Fishes were caught in two different spawning areas of the Strait of Sicily, characterized by different environmental conditions, including a coastal upwelling with a lower temperature (Adventure Bank; 20.22 ± 0.38°C) and a thermohaline front with a higher temperature (Maltese Bank 23.10 ± 0.25°C). The results showed that the two groups of larvae, in their early life, had similar nutritional status. However, compared with the samples from the Maltese Bank, the specimens collected in the Adventure Bank area exhibited higher alkaline phosphatase activity, lower concentrations of aspartate plus asparagine, threonine, and arginine but a higher concentration of leucine, highlighting different patterns of amino acid metabolism. Collectively, these results indicated that AAC analysis could represent an additional valid tool to evaluate the link between physiological responses and environmental conditions at early life stages.

Analysis of sugars and amino acids in aphid honeydew by hydrophilic interaction liquid chromatography – Mass spectrometry

Past analyses of sugar and amino acid composition of aphid honeydews have been completed using diverse instrumentation. Here we report the use of hydrophilic interaction liquid chromatography (HILIC) coupled with a triple quadrupole mass spectrometric (MS/MS) detector for the analysis of seven saccharides (xylose, fructose, glucose, sucrose, trehalose, melezitose and raffinose) and five amino acids (glutamic acid, glutamine, aspartic acid, serine, and asparagine). Limits of quantitation ranged from 0.05 mg/L (melezitose) to 1.0 mg/L (fructose) for sugars and from 0.10 mg/L (glutamic acid) to 3.66 mg/L (asparagine) for amino acids. Sample preparation was fast and simple, requiring only the washing of foils used to collect aphid honeydew with hot (80 °C) water and sonication of samples prior to HILIC/MS/MS analysis for both classes of analytes. No analyte derivatization was required and excellent chromatographic characteristics were observed. For those studying honeydew-mediated interactions in the field, this technique allows for rapid characterization of ecologically important amino acids and sugars.•Composition of seven saccharides in Aphis asclepiadis honeydew including xylose, fructose, glucose, sucrose, trehalose, melezitose,and raffinose, and five standard amino acids including glutamic acid, glutamine, aspartic acid, serine, and asparagine, were analyzed using hydrophilic interaction liquid chromatography-mass spectrometry.•All polar analytes were analyzed without derivatization using HILIC-MS with chromatographic run times of 7 min (sugars) and 10 min (amino acids).

Study of Acetolactate Synthase and its Mechanism of Inhibition by Sulfonylurea Active Ingredients: Amidosulfuron, Nicosulfuron, Cyclosulfuron – In-silico Approach

Acetolactate synthase (ALS) or Acetohydroxyacid synthase (AHAS) catalyzes the first step in the synthesis of the branched-chain amino acids i.e., valine (2-amino-3-methylbutanoic acid), leucine (2-amino-4-methylpentanoic acid), and isoleucine (2-amino-3-methylpentanoic acid), in plants, bacteria, algae and fungi but not in humans. AHAS is the main target enzyme for sulfonylurea active ingredients; Amidosulfuron, Nicosulfuron and Cyclosulfuron those assist in lowering branched-chain amino acid synthesis through inhibition to form the complex of Lactyl-ThDP(TDL) to ALS with great practical importance. Amino acid composition, evolutionary and sequence analysis of the ALS protein from Arabidopsis thaliana and its homologous were systematically studied. Composition analysis reveals that ALS is a soluble protein. Moreover, the phylogenetic tree showed different clusters based on the source organism and multiple sequence alignment depicts conservative nature in amino acid residues. Furthermore, molecular docking has been conducted to study the interactions between ALS of Arabidopsis thaliana and TDL in presence/absence of the active ingredients of sulfonylurea herbicide groups. Molecular docking studies confirm active ingredients are effective to inhibit the binding of TDL to ALS. Our obtained results can be very useful to study specific protein interactions along with developing new herbicides using computational methods.

Physical and oxidative stability of chicken oil-in-water emulsion stabilized by chicken protein hydrolysates.

The emulsifying and antioxidant properties of chicken protein hydrolysates for the physical and oxidative stabilization of chicken oil‐in‐water emulsion were investigated. The chicken protein pepsin hydrolysates obtained at reaction temperature of 33℃, 1.8% enzyme addition, liquid–solid ratio of 5:1, and reaction time of 4h, showed the DPPH radical scavenging rate of 92.12% and emulsion stability index of 0.07. The hydrolysate exerted significantly improved antioxidant activity and emulsion ability compared to the native chicken protein. The amino acid composition analysis indicated that the contents of hydrophobic amino acids including tyrosine, phenylalanine, and tryptophan were increased after hydrolysis, which contributed to the higher hydrophobicity and antioxidant activity of chicken hydrolysates. The results suggested that the chicken protein hydrolysates could be used as an alternative protein emulsifier for the production of oxidatively stable chicken oil‐in‐water emulsion.

Angiotensin converting enzyme inhibitory peptide fractions from Tibet wild peach kernel protein hydrolysates

In this study, the peach kernel proteins were extracted and treated with alkaline proteinase to generate peach kernel protein hydrolysate (PKH), which showed angiotensin converting enzyme (ACE) inhibition activity. The hydrolysate was separated into four fractions and their anti-ACE activities were investigated. Our results showed that all PKHs had anti-ACE activity, and the lowest molecular weight fraction PKH4 had the highest ACE inhibitory activity. Lineweaver–Burk plots illustrated that the inhibition types of PKH3 and PKH4 were non-competitive. The Ki of PKH4 was lower than Ki of PKH3; suggesting PKH4 had high affinity to ACE. Amino acid composition analysis showed that the best anti-ACE peptide PKH4 possessed high levels of hydrophobic amino acids, branched-chain amino acids, and aromatic amino acids. In summary, our findings demonstrated that high anti-ACE activity is negatively related to the size of the PKHs and possibly the composition of amino acids, and the PKH4 was the best ACE inhibitor. Further, peach kernel peptides can be developed as a functional food for patients with hypertension.

Feasibility of synthesizing γ-[Glu](n≥1)-Gln using high solid concentrations and glutaminase from Bacillus amyloliquefaciens as the catalyst

Effects of using high solid concentrations on the synthesis of γ-[Glu](n≥1)-Gln catalyzed by glutaminase from Bacillus amyloliquefaciens were examined in this study. An increment in solid concentration from 10% to 50% (w/w) increased the extent of synthesis of γ-[Glu](n≥1)-Gln, based on the analyses of amino acid composition and amino nitrogen content. Size-exclusion high-performance liquid chromatography analysis revealed an increase in molecular mass of γ-[Glu](n≥1)-Gln resulting from the increase of solid concentration from 10% to 50% (w/w). UPLC-Q-TOF-MS/MS analysis showed that the enzymatic reaction mixtures post γ-glutamyl transpeptidation contained γ-Glu-Gln, γ-Glu-Glu-Gln, γ-Glu-Glu-Glu-Gln, γ-Glu-Glu-Glu-Glu-Gln and γ-Glu-Glu-Glu-Glu-Glu-Gln. The intensity of each γ-[Glu](n=1,2,3,4,5)-Gln produced at a solid concentration of 50% (w/w) was higher than that at 10% (w/w). These findings indicated the potential of such an energy- and water-efficient approach for synthesizing γ-[Glu](n≥1)-Gln at high solid concentrations.

Synthesis and preliminary evaluation of neoglycopolymers carrying multivalent N-glycopeptide units

In the present study, for the discovery of uncharacterized glycan-binding receptors or lectin-like receptors in plants, we developed neoglycopolymers to which three types of N-glycopeptides are conjugated; the first with plant complex type N-glycan (M3FX), the second with high-mannose type N-glycan (M8), and the third with animal complex type N-glycan (NeuAc2Gal2GN2M3). Three types of Asn-oligosaccharide (Asn-M3FX, Asn-M8, or Asn-NeuAc2Gal2GN2M3) were prepared from storage glycoproteins of Ginkgo biloba seeds, Vigna angularis seeds, and egg yolk glycopeptides from actinase digests of each glycoproteins or glycopeptide. Neoglycopolymers were synthesized such that the α-amino groups of Asn-oligosaccharide were coupled to the carboxyl groups of poly-γ-L-glutamic acid (γ-L-PGA) with 4-(4,6-Dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride n-hydrate (DMT-MM). The resulting neoglycopolymers were purified through a combination of gel-filtration and reverse-phase HPLC. The incorporation of N-glycans into γ-L-PGA (mol%) was estimated through amino acid composition analysis after acid hydrolysis. The incorporation rates of Asn-M3FX, Asn-M8, and Asn-NeuAc2Gal2GN2M3 into γ-L-PGA were 15.4%, 8.6%, and 11.1%, indicating that nearly 890, 500, and 640 molecules of N-glycans were conjugated with γ-L-PGA, respectively. Furthermore, we confirmed that the neoglycopolymer carrying the multivalent high-mannose type N-glycans is a useful tool for rapid purification of mannose-binding protein, Concanavalin A, from jack bean extract.

Purification and sweetness properties of egg white lysozymes from Indonesian local poultry of ayam kampung and Cihateup duck

Egg white lysozyme (EWL) is known to elicit a sweet response with the sweetness threshold was higher than sucrose. It, however, remains to be examined if EWL from different poultry exhibited similar sweetness properties. This study aims to characterize sweetness properties of EWL from Indonesian local poultry of ayam kampung (AK-EWL) and Cihateup duck (CD-EWL). Accordingly, AK-EWL and CD-EWL were purified from the egg white using an Amberlite FPC3500 resin. As a control, EWL of non-indonesian laying hen (hen-EWL) was also purified using the same method. Enzymatic activities of hen-EWL, AK-EWL and CD-EWL were found to be different, whereby AK-EWL exhibited the highest activity. The result confirmed that hen-EWL, AK-EWL and CD-EWL exhibited sweet responses with the threshold of 8.45, 10.39, and 7.66 µM, respectively. Amino acid composition analysis revealed that these three EWLs are dominated by non-polar amino acids. Nevertheless, the ratio of positively and negatively charged residues among them varied which might account for the differences on the sweetness threshold of hen-EWL, AK-EWL and CD-EWL. Further, pasteurization and sterilization were found to have significantly reduced the enzymatic activity, sweetness and structures of hen-EWL, AK-EWL and CD-EWL This suggested that the practical application of hen-EWL, AK-EWL and CD-EWL as sweeteners should not involve any heating treatments that might reduce their sweetness levels.

Effects of Hydrolyzed Animal Protein on the Enhancement of Saltiness and Quality Characteristics of White Pan Bread

The saltiness enhancer effect of hydrolyzed animal protein (HAP) and the quality characteristics of white pan bread were evaluated through physicochemical and sensory analyses. HAP was efficiently hydrolyzed using commercial enzymes (0.20% w/w Alcalase® 2.4 L FG and 1.00% w/w FlavourzymeTM 500 MG) under high pressurization (100 MPa) and confirmed by molecular weight distribution and amino acid composition analysis. Most HAP molecules (94.18%) comprised low molecular weight peptides < 1300 Da, or free amino acids. Amino acid composition analysis of HAP also detected highly increased glutamic acid concentration. Compared with the control, the fermentation rates of doughs and loaf volumes of breads were maintained over 89.27% and 100%, respectively, with an exception of samples containing 11.70 HAP. In sensory testing, HAP concentration increases led to concomitant saltiness increases. These results suggest promising technological options for reducing salt intake from white pan bread consumption by utilizing HAP, although further study is necessary to improve bread quality.

Cloning and characterization of a thermostable glutathione reductase from a psychrophilic Arctic bacterium Sphingomonas sp.

Glutathione reductase is an important oxidoreductase that helps maintain redox homeostasis by catalyzing the conversion of glutathione disulfide to glutathione using NADPH as a cofactor. In this study, we cloned and characterized a glutathione reductase (hereafter referred to as SpGR) from Sphingomonas sp. PAMC 26621, an Arctic bacterium. SpGR comprises 449 amino acids, and functions as a dimer. Surprisingly, SpGR exhibits characteristics of thermophilic enzymes, showing optimum activity at 60°C and thermal stability up to 70°C with ∼50% residual activity at 70°C for 2 h. The amino acid composition analysis of SpGR showed a 1.9-fold higher Arg content (6%) and a 2.7-fold lower Lys/Arg ratio (0.75) compared to the Arg content (3.15%) and the Lys/Arg ratio (2.01) of known psychrophilic glutathione reductases. SpGR also exhibits its activity at 4°C, and circular dichroism and fluorescence spectroscopy results indicate that SpGR maintains its secondary and tertiary structures within the temperature range of 4-70°C. Taken together, the results of this study indicate that despite its origin from a psychrophilic bacterium, SpGR has high thermal stability. Our study provides an insight into the role of glutathione reductase in maintaining the reducing power of an Arctic bacterium in a broad range of temperatures.

The molecular mass and isoelectric point of plant proteomes

BackgroundCell contain diverse array of proteins with different molecular weight and isoelectric point (pI). The molecular weight and pI of protein play important role in determining the molecular biochemical function. Therefore, it was important to understand the detail regarding the molecular weight and pI of the plant proteins.ResultsA proteome-wide analysis of plant proteomes from 145 species revealed a pI range of 1.99 (epsin) to 13.96 (hypothetical protein). The spectrum of molecular mass of the plant proteins varied from 0.54 to 2236.8 kDa. A putative Type-I polyketide synthase (22244 amino acids) in Volvox carteri was found to be the largest protein in the plant kingdom. However, Type-I polyketide synthase was not found in higher plant species. Titin (806.46 kDa) and misin/midasin (730.02 kDa) were the largest proteins identified in higher plant species. The pI and molecular weight of the plant proteins showed a trimodal distribution. An acidic pI (56.44% of proteins) was found to be predominant over a basic pI (43.34% of proteins) and the abundance of acidic pI proteins was higher in unicellular algae species relative to multicellular higher plants. In contrast, the seaweed, Porphyra umbilicalis, possesses a higher proportion of basic pI proteins (70.09%). Plant proteomes were also found to contain selenocysteine (Sec), amino acid that was found only in lower eukaryotic aquatic plant lineage. Amino acid composition analysis showed Leu was high and Trp was low abundant amino acids in the plant proteome. Additionally, the plant proteomes also possess ambiguous amino acids Xaa (unknown), Asx (asparagine or aspartic acid), Glx (glutamine or glutamic acid), and Xle (leucine or isoleucine) as well.ConclusionThe diverse molecular weight and isoelectric point range of plant proteome will be helpful to understand their biochemical and functional aspects. The presence of selenocysteine proteins in lower eukaryotic organism is of interest and their expression in higher plant system can help us to understand their functional role.