共查询到20条相似文献,搜索用时 15 毫秒
1.
Verónica Beatriz Rajal Alicia Graciela Cid Guillermo Ellenrieder Carlos Mario Cuevas 《World journal of microbiology & biotechnology》2009,25(6):1025-1033
Penicillium ulaiense is a post-harvest pathogenic fungus that attacks citrus fruits. The objective of this work was to study this microorganism
as an α-l-rhamnosidase producer and to characterize it from P. ulaiense. The enzyme under study is used for different applications in food and beverage industries. α-l-Rhamnosidase was produced in a stirred-batch reactor using rhamnose as the main carbon source. The kinetic parameters for
the growth of the fungi and for the enzyme production were calculated from the experimental values. A method for partial purification,
including (NH4)2SO4 precipitation, incubation at pH 12 and DEAE-sepharose chromatography yielded an enzyme with very low β-glucosidase activity.
The pH and temperature optima were 5.0 and 60°C, respectively. The Michaelis–Menten constants for the hydrolysis of p-nitrophenyl-α-l-rhamnoside were V
max = 26 ± 4 IU ml−1 and K
m
= 11 ± 2 mM. The enzyme showed good thermostability up to 60°C and good operational stability in white wine. Co2+ affected positively the activity; EDTA, Mn2+, Mg2+, dithiotreitol and Cu2+ reduced the activity by different amounts, and Hg2+ completely inhibited the enzyme. The enzyme showed more activity on p-nitrophenyl-α-l-rhamnoside than on naringin. According to these results, this enzyme has potential for use in the food and pharmacy industries
since P. ulaiense does not produce mycotoxins. 相似文献
2.
β-N-Methylamino-l-alanine (BMAA), a non-proteinogenic amino acid, has been detected in a range of cyanobacteria, including terrestrial, aquatic,
free living and endosymbiotic species. The widespread occurrence of cyanobacteria in the environment raises concerns regarding
the ecological and toxicological impact of BMAA, and consequently, studies have focussed extensively on the toxicity and environmental
impact of BMAA, while no research has addressed the ecophysiological or metabolic role of the compound in cyanobacteria. In
this study, both the uptake of exogenous BMAA by and the effect of exogenous BMAA on the growth of Synechocystis PCC6803 were investigated. BMAA was rapidly taken up by the non-diazotrophic cyanobacterium Synechocystis PCC6803 in a concentration dependent manner. The presence of exogenous BMAA resulted in a substantial and concentration-dependent
decrease in cell growth and the substantial loss of photosynthetic pigmentation. Similar effects were seen in the presence
of the non-proteinogenic amino acid, 2,4-diaminobutyric acid but to a lesser degree than that of BMAA. The effects were reversed
when light was decreased from 16 to 10 μmol m−2 s−1. Control cultures grown in the presence of l-arginine, l-asparagine, l-glutamate and glycine showed normal or slightly increased growth with no change in pigmentation. The decrease in growth rate
coupled to bleaching indicates that BMAA may induce chlorosis in the presence of adequate photosynthetic radiation suggesting
a connection between BMAA and the induction of conditions, such as nitrogen or sulphur depletion, that result in growth arrest
and the induction of chlorosis. 相似文献
3.
Poly(ε-l-lysine) (ε-PL) is a naturally occurring poly(amino acid) characterized by a unique structure linking ε-amino and carboxyl
groups of l-lysine. Due to its various functions and its biodegradability and non-toxicity, the ε-PL polymer has attracted increasing
attention in recent years. ε-PL is frequently found in various strains of Streptomyces sp. This review gives an up-to-date overview regarding the biosynthesis of ε-PL focussing mainly on results obtained from
ten newly isolated producer strains, using the two-stage culture method of cell growth and ε-PL production cultures. The production
of nearly monodispersed ε-PL is covered together with the development of ε-PL specific hydrolases and the release of synthesized
ε-PL into the culture broth. From these results, coupled with the termination of polymerization through nucleophilic chain
transfer, the biosynthetic mechanism of the polymer is discussed. 相似文献
4.
Soria F Ellenrieder G Oliveira GB Cabrera M Carvalho LB 《Applied microbiology and biotechnology》2012,93(3):1127-1134
α-l-Rhamnosidase from Aspergillus terreus was covalently immobilized on the following ferromagnetic supports: polyethylene terephthalate (Dacron-hydrazide), polysiloxane/polyvinyl
alcohol (POS/PVA), and chitosan. The powdered supports were magnetized by thermal coprecipitation method using ferric and
ferrous chlorides, and the immobilization was carried out via glutaraldehyde. The activity of the Dacron-hydrazide (0.53 nkat/μg
of protein) and POS/PVA (0.59 nkat/μg of protein) immobilized enzyme was significantly higher than that found for the chitosan
derivative (0.06 nkat/μg of protein). The activity–pH and activity–temperature profiles for all immobilized enzymes did not
show difference compared to the free enzyme, except the chitosan derivative that presented higher maximum temperature at 65 °C.
The Dacron-hydrazide derivative thermal stability showed a similar behavior of the free enzyme in the temperature range of
40–70 °C. The POS/PVA and chitosan derivatives were stable up to 60 °C, but were completely inactivated at 70 °C. The activity
of the preparations did not appreciably decrease after ten successive reuses. Apparent K
m of α-l-rhamnosidase immobilized on magnetized Dacron-hydrazide (1.05 ± 0.22 mM), POS/PVA (0.57 ± 0.09 mM), and chitosan (1.78 ± 0.24 mM)
were higher than that estimated for the soluble enzyme (0.30 ± 0.03 mM). The Dacron-hydrazide enzyme derivative showed better
performance than the free enzyme to hydrolyze 0.3% narigin (91% and 73% after 1 h, respectively) and synthesize rhamnosides
(0.116 and 0.014 mg narirutin after 1 h, respectively). 相似文献
5.
de Wet BJ Matthew MK Storbeck KH van Zyl WH Prior BA 《Applied microbiology and biotechnology》2008,77(5):975-983
A glycosyl hydrolase family 54 (GH54) α-l-arabinofuranosidase gene (abfA) of Aureobasidium pullulans was amplified by polymerase chain reaction from genomic DNA and a 498-amino-acid open reading frame deduced from the DNA
sequence. Modeling of the highly conserved A. pullulans AbfA protein sequence on the crystal structure of Aspergillus kawachii AkabfB showed that the catalytic amino acid arrangement and overall structure were highly similar including the N-terminal
catalytic and C-terminal arabinose binding domains. The abfA gene was expressed in Saccharomyces cerevisiae, and the heterologous enzyme was purified. The protein was monomeric, migrating at 49 kDa on sodium dodecyl sulfate-polyacrylamide
gel electrophoresis and eluting at 36 kDa upon gel filtration. AbfA showed maximal activity at 55°C and between pH 3.5 and
pH 4. The enzyme had a K
m value for p-nitrophenyl-α-l-arabinofuranoside of 3.7 mM and a V
max of 34.8 μmol min−1 mg protein−1. Arabinose acted as a noncompetitive inhibitor with a K
i of 38.4 mM. The enzyme released arabinose from maize fiber, oat spelt arabinoxylan, and wheat arabinoxylan, but not from
larch wood arabinogalactan or α-1,5-debranched arabinan. AbfA displayed low activity against α-1,5-l-arabino-oligosaccharides. The enzyme acted synergistically with endo-β-1,4-xylanase in the breakdown of wheat arabinoxylan. Binding of AbfA to xylan from several sources confirmed the presence
of a functional carbohydrate-binding module.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
6.
l-β-Haloalanines are physiologically active unnatural amino acids and they are useful intermediates for the synthesis of natural
and unnatural amino acids, S-linked glycopeptides, and lanthionines. In general l-β-haloalanines were prepared predominantly from l-serine via functional group transformation. Here we reported an alternative approach for the preparation of l-β-haloalanines via halogenation of protected l-cysteine esters which was obtained from l-cysteine or l-cystine, respectively. The mercapto group of protected l-cysteine esters was efficiently transformed to halo groups by triphenylphosphine/N-halosuccinimides. It has been proved to be a versatile desulfurization strategy via this functional group transformation. 相似文献
7.
Flax seed mucilage (FM) contains a mixture of highly doubly substituted arabinoxylan as well as rhamnogalacturonan I with
unusual side group substitutions. Treatment of FM with a GH11 Bacillus subtilis XynA endo 1,4-β-xylanase (BsX) gave limited formation of reducing ends but when BsX and FM were incubated together on different
wheat arabinoxylan substrates and birchwood xylan, significant amounts of xylose were released. Moreover, arabinose was released
from both water-extractable and water-unextractable wheat arabinoxylan. Since no xylose or arabinose was released by BsX addition
alone on these substrates, nor without FM or BsX addition, the results indicate the presence of endogenous β-d-xylosidase and α-l-arabinofuranosidase activities in FM. FM also exhibited activity on both p-nitrophenyl α-l-arabinofuranoside (pNPA) and p-nitrophenyl β-d-xylopyranoside (pNPX). Based on K
M
values, the FM enzyme activities had a higher affinity for pNPX (K
M
2 mM) than for pNPA (K
M
20 mM). 相似文献
8.
A bacterial strain, MAK-2, was isolated as a producer of α-l-rhamnosidase from a soil sample of Dehradoon, India. The strain was identified based on morphology, physiological tests and
16S rDNA analysis. The phylogenetic analysis based on the 16S rDNA sequence, identified the isolate as Staphylococcus xylosus, a non-pathogenic member of CNS (coagulase-negative staphylococci) family. The strain was capable of producing α-l-rhamnosidase by hydrolysing flavonoids thus confirming potential application in the citrus-processing industry. 相似文献
9.
Cobucci-Ponzano B Conte F Rossi M Moracci M 《Extremophiles : life under extreme conditions》2008,12(1):61-68
Glycoside hydrolases form hyperthermophilic archaea are interesting model systems for the study of catalysis at high temperatures
and, at the moment, their detailed enzymological characterization is the only approach to define their role in vivo. Family
29 of glycoside hydrolases classification groups α-l-fucosidases involved in a variety of biological events in Bacteria and Eukarya. In Archaea the first α-l-fucosidase was identified in Sulfolobus solfataricus as interrupted gene expressed by programmed −1 frameshifting. In this review, we describe the identification of the catalytic
residues of the archaeal enzyme, by means of the chemical rescue strategy. The intrinsic stability of the hyperthermophilic
enzyme allowed the use of this method, which resulted of general applicability for β and α glycoside hydrolases. In addition,
the presence in the active site of the archaeal enzyme of a triad of catalytic residues is a rather uncommon feature among
the glycoside hydrolases and suggested that in family 29 slightly different catalytic machineries coexist. 相似文献
10.
Kapil Tahlan Marcus A. Moore Susan E. Jensen 《Journal of industrial microbiology & biotechnology》2017,44(4-5):517-524
The δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine (ACV) tripeptide is the first dedicated intermediate in the biosynthetic pathway leading to the penicillin and cephalosporin classes of β-lactam natural products in bacteria and fungi. It is synthesized nonribosomally by the ACV synthetase (ACVS) enzyme, which has been purified and partially characterized from many sources. Due to its large size and instability, many details regarding the reaction mechanism of ACVS are still not fully understood. In this review we discuss the chronology and associated methodology that led to the discovery of ACVS, some of the main findings regarding its activities, and some recent/current studies being conducted on the enzyme. In addition, we conclude with perspectives on what can be done to increase our understating of this very important protein in the future. 相似文献
11.
Setlow B Cabrera-Hernandez A Cabrera-Martinez RM Setlow P 《Archives of microbiology》2004,181(1):60-67
Four aryl-phospho--d-glucosidases were identified in Bacillus subtilis by using 4-methylumbelliferyl-phospho--d-glucopyranoside as a substrate. Two of these enzymes are the products of the bglA and bglH genes, previously suggested to encode aryl-phospho--d-glucosidases, while the other enzymes are encoded by the yckE and ydhP genes. Together, these four genes account for >99.9% of the glucosidase activity in B. subtilis on aryl-phospho--d-glucosides. yckE was expressed at a low and constant level during growth, sporulation, and spore germination, and was not induced by aryl--d-glucosides. ydhP was also not induced by aryl--d-glucosides. However, while ydhP was expressed at only a very low level in exponential-phase cells and germinating spores, this gene was expressed at a higher levels upon entry into the stationary phase of growth. Strains lacking yckE or ydhP exhibited no defects in growth, sporulation, or spore germination or in growth on aryl--d-glucosides. However, a strain lacking bglA, bglH and yckE grew poorly if at all on aryl--d-glucosides as the sole carbon source.Abbreviations
MU
4-Methylumbelliferone
-
MUG
4-Methylumbelliferyl--d-glucopyranoside
-
MUGal
4-Methylumbelliferyl--d-galactopyranoside
-
MUG-P
4-Methylumbelliferyl--d-glucopyranoside-6-phosphate 相似文献
12.
Theodoros Goulas Athanasios Goulas George Tzortzis Glenn R. Gibson 《Applied microbiology and biotechnology》2009,84(5):899-907
This paper deals with two aspects tightly related to the enzymatic characteristics and expression of four β-galactosidases (BbgI, BbgII, BbgIII and BbgIV) from Bifidobacterium bifidum NCIMB41171. The growth patterns of this strain indicated a preference towards complex (i.e. lactose, galactooligosaccharides
(GOSs)) rather than simple carbohydrates (i.e. glucose and galactose) and a collaborative action and synergistic relation
of more than one β-galactosidase isoenzyme for either lactose or GOS hydrolysis and subsequent assimilation. Native polyacrylamide gel electrophoresis
analysis of protein extracts from cells growing on different carbohydrates (i.e. glucose, lactose or GOS) indicated that two
lactose hydrolysing enzymes (BbgI and BbgIII) and one GOS hydrolysing enzyme (BbgII) were constitutively expressed, whereas
a fourth lactose hydrolysing enzyme (BbgIV) was induced in the presence of lactose or different GOS fractions. Furthermore,
the β-galactosidase expression profiles of B. bifidum cells and the transgalactosylating properties of each individual isoenzyme, with lactose as substrate, clearly indicated
that mainly three isoenzymes (BbgI, BbgIII and BbgIV) are implicated in GOS synthesis when whole B. bifidum cells are utilised. Two of the isoenzymes (BbgI and BbgIV) proved to have better transgalactosylating properties giving yields
ranging from 42% to 47% whereas the rest (BbgI and BbgIII) showed lower yields (15% and 29%, respectively). 相似文献
13.
Xiufeng Shi Chuanyou Chang Shenxi Ma Yibing Cheng Jun Zhang Qiang Gao 《Journal of industrial microbiology & biotechnology》2017,44(4-5):697-704
This work investigated the efficient bioconversion process of l-glutamate to GABA by Lactobacillus brevis TCCC 13007 resting cells. The optimal bioconversion system was composed of 50 g/L 48 h cultivated wet resting cells, 0.1 mM pyridoxal phosphate in glutamate-containing 0.6 M citrate buffer (pH 4.5) and performed at 45 °C and 180 rpm. By 10 h bioconversion at the ratio of 80 g/L l-glutamic acid to 240 g/L monosodium glutamate, the final titer of GABA reached 201.18 g/L at the molar bioconversion ratio of 99.4 %. This process presents a potential for industrial and commercial applications and also offers a promising feasibility of continuous GABA production coupled with fermentation. Besides, the built kinetics model revealed that the optimum operating conditions were 45 °C and pH 4.5, and the bioconversion kinetics at low ranges of substrate concentration (0 < S < 80 g/L) was assumed to follow the classical Michaelis–Menten equation. 相似文献
14.
K. Škrlep M. Bergant G. M. De Winter B. Bohanec J. Žel R. Verpoorte F. Van Iren M. Camloh 《Biologia Plantarum》2008,52(2):329-333
Different lines of cell suspension cultures of Taxus × media Rehd. and Taxus floridana Nutt. were cryopreserved with a two-step freezing method using a simple and inexpensive freezing container instead of a programmable
freezer. Four to seven days old suspension cell cultures were precultured in growth medium supplemented with 0.5 M mannitol
for 2 d. The medium was then replaced with cryoprotectant solution (1 M sucrose, 0.5 M glycerol and 0.5 M dimethylsulfoxide)
and the cells incubated on ice for 1 h. Before being plunged into liquid nitrogen, cells were frozen with a cooling rate of
approximately −1 °C per min to −80 °C. The highest post-thaw cell viability was 90 %. The recovery was line dependent. The
cryopreservation procedure did not alter the nuclear DNA content of the cell lines. The results indicate that cryopreservation
of Taxus cell suspension cultures using inexpensive freezing container is possible. 相似文献
15.
Rojas NL Voget CE Hours RA Cavalitto SF 《Journal of industrial microbiology & biotechnology》2011,38(9):1515-1522
Rhamnosidases are enzymes that catalyze the hydrolysis of terminal nonreducing L-rhamnose for the bioconversion of natural or synthetic rhamnosides. They are of great significance in the current biotechnological
area, with applications in food and pharmaceutical industrial processes. In this study we isolated and characterized a novel
alkaline rhamnosidase from Acrostalagmus luteo albus, an alkali-tolerant soil fungus from Argentina. We also present an efficient, simple, and inexpensive method for purifying
the A. luteo albus rhamnosidase and describe the characteristics of the purified enzyme. In the presence of rhamnose as the sole carbon source,
this fungus produces a rhamnosidase with a molecular weight of 109 kDa and a pI value of 4.6, as determined by SDS–PAGE and
analytical isoelectric focusing, respectively. This enzyme was purified to homogeneity by chromatographic and electrophoretic
techniques. Using p-nitrofenil-α-L-rhamnopiranoside as substrate, the enzyme activity showed pH and temperature optima of 8.0 and 55°C, respectively. The enzyme
exhibited Michaelis–Menten kinetics, with K
M and V
max values of 3.38 mmol l−1 and 68.5 mmol l−1 min−1, respectively. Neither divalent cations such as Ca2+, Mg2+, Mn2+, and Co2+ nor reducing agents such as β-mercaptoethanol and dithiothreitol showed any effect on enzyme activity, whereas this activity
was completely inhibited by Zn2+ at a concentration of 0.2 mM. This enzyme showed the capacity to hydrolyze some natural rhamnoglucosides such as hesperidin,
naringin and quercitrin under alkaline conditions. Based on these results, and mainly due to the high activity of the A. luteo albus rhamnosidase under alkaline conditions, this enzyme should be considered a potential new biocatalyst for industrial applications. 相似文献
16.
17.
Summary The catalytic amino acid residues of the extracellular β-D-glucosidase (β-D-glucoside glucohydrolase, EC 3.2.1.21) from Aspergillus carbonarius were investigated. The pH dependence curves gave apparent pK values of 2.8 and 5.93 for the free enzyme, and 2.24 and 6.14 for the enzyme–substrate complex using p-nitrophenyl-β-D-glucoside as substrate. Carbodiimide- and Woodward reagent K-mediated chemical modifications suggested that a carboxylate residue, located in the active centre, was fundamental in the catalysis. The pH dependence of inactivation revealed the involvement of a group with pK value of 4.61 in the modification reaction, proving that a carboxylate residue was modified. The A. carbonarius β-glucosidase was irreversibly inactivated by N-bromoacetyl-β-D-glucopyranosylamine. The active site specificity of the inactivation was proved by using the competitive inhibitor p-nitrophenyl-1-thio-β-D-glucopyranoside. pH Dependence studies of inactivation revealed that modification by N-bromoacetyl-β-D-glucopyranosylamine could be directed toward the carboxylate group acting as the catalytic nucleophile, as in the case of the carbodiimide and Woodward reagent K modifications. 相似文献
18.
Thermomonospora fusca produced a relatively high level of alpha-L-arabinofuranosidase when growing on oat spelt xylan as the main carbon and energy source. The enzyme exhibited maximum relative activity (0.136 U/g protein) at pH 9.0 with 54 and 55% activity remaining at pH of 4.5 and 11.0, respectively. The apparent Km value for the crude alpha-L-arabinofuranosidase preparation was 180 mumol/L 4-nitrophenyl alpha-L-arabinofuranoside; the upsilon lim value was the release of 40 mumol/L 4-nitrophenol per min. Enzyme activity was eluted as a single peak (HPLC gel filtration chromatography) corresponding to molar mass of approximately 92 kDa. Native electrophoresis of crude cell lysate confirmed the presence of a single active intracellular alpha-L-arabinofuranosidase component. SDS-PAGE of this enzyme, developed as zymogram, did not demonstrate any activity; denaturing gel was stained and a protein band of relative molar mass of 46 kDa was revealed. Isoelectric focusing of a purified alpha-L-arabinofuranosidase yielded a single protein band for the corresponding activity zone with pI 7.9. The enzyme was purified approximately 21-fold the mean overall yield was about 16%. 相似文献
19.
Scheibner M Hülsdau B Zelena K Nimtz M de Boer L Berger RG Zorn H 《Applied microbiology and biotechnology》2008,77(6):1241-1250
Two extracellular enzymes (MsP1 and MsP2) capable of efficient β-carotene degradation were purified from culture supernatants
of the basidiomycete Marasmius scorodonius (garlic mushroom). Under native conditions, the enzymes exhibited molecular masses of ~150 and ~120 kDa, respectively. SDS-PAGE
and mass spectrometric data suggested a composition of two identical subunits for both enzymes. Biochemical characterisation
of the purified proteins showed isoelectric points of 3.7 and 3.5, and the presence of heme groups in the active enzymes.
Partial amino acid sequences were derived from N-terminal Edman degradation and from mass spectrometric ab initio sequencing of internal peptides. cDNAs of 1,604 to 1,923 bp, containing open reading frames (ORF) of 508 to 513 amino acids,
respectively, were cloned from a cDNA library of M. scorodonius. These data suggest glycosylation degrees of ~23% for MsP1 and 8% for MsP2. Databank homology searches revealed sequence
homologies of MsP1 and MsP2 to unusual peroxidases of the fungi Thanatephorus cucumeris (DyP) and Termitomyces albuminosus (TAP). 相似文献
20.
A lectin present in seeds of Clitoria ternatea agglutinated trypsin-treated human B erythrocytes. The sugar specificity assay indicated that lectin belongs to Gal/Gal NAc-specific
group. Hence the lectin, designated C. ternatea agglutinin (CTA), was purified by the combination of acetic acid precipitation, salt fractionation and affinity chromatography.
HPLC gel filtration, SDS-polyacrylamide gel electrophoresis and mass spectrometry indicated that the native lectin is composed
of two identical subunits of molecular weight 34.7 kDa associated by non covalent bonds. The N-terminal sequence of CTA shared
homology with Glycine max and Pisum sativum. Complete sequence was also found to be homologous to S-64 protein of Glycine max, suggesting that CTA probably exhibits both hemagglutination and probably sugar uptake activity. The carbohydrate binding
specificity of the lectin was investigated by quantitative turbidity measurements, and percent inhibition assays. Based on
these assays, we conclude that CTA binds β-d-galactosides, and also may has an extended specificity towards non-reducing terminal Neu5Acα2,6Gal. 相似文献