scholarly journals Cloning and Characterization of a Novel Alginate Lyase from Paenibacillus sp. LJ-23

Marine Drugs ◽  
2022 ◽  
Vol 20 (1) ◽  
pp. 66
Author(s):  
Mingpeng Wang ◽  
Lei Chen ◽  
Zhengyu Lou ◽  
Xueting Yuan ◽  
Guiping Pan ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Water Solubility ◽  
Degradation Products ◽  
Three Dimensional ◽  
Alginate Lyase ◽  
Dimensional Structure ◽  
Amino Acid Residues ◽  
Alginate Oligosaccharides ◽  
Paenibacillus Sp ◽  
Comprehensive Health

As a low molecular weight alginate, alginate oligosaccharides (AOS) exhibit improved water solubility, better bioavailability, and comprehensive health benefits. In addition, their biocompatibility, biodegradability, non-toxicity, non-immunogenicity, and gelling capability make them an excellent biomaterial with a dual curative effect when applied in a drug delivery system. In this paper, a novel alginate lyase, Algpt, was cloned and characterized from a marine bacterium, Paenibacillus sp. LJ-23. The purified enzyme was composed of 387 amino acid residues, and had a molecular weight of 42.8 kDa. The optimal pH of Algpt was 7.0 and the optimal temperature was 45 °C. The analysis of the conserved domain and the prediction of the three-dimensional structure indicated that Algpt was a novel alginate lyase. The dominant degradation products of Algpt on alginate were AOS dimer to octamer, depending on the incubation time, which demonstrated that Algpt degraded alginate in an endolytic manner. In addition, Algpt was a salt-independent and thermo-tolerant alginate lyase. Its high stability and wide adaptability endow Algpt with great application potential for the efficient preparation of AOS with different sizes and AOS-based products.

scholarly journals Functional Analysis of the Mycobacterium tuberculosis FAD-Dependent Thymidylate Synthase, ThyX, Reveals New Amino Acid Residues Contributing to an Extended ThyX Motif

2008 ◽  
Vol 190 (6) ◽  
pp. 2056-2064 ◽  
Author(s):  
Jonathan E. Ulmer ◽  
Yap Boum ◽  
Christopher D. Thouvenel ◽  
Hannu Myllykallio ◽  
Carol Hopkins Sibley
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Thymidylate Synthase ◽  
Three Dimensional ◽  
Pyrimidine Ring ◽  
Dimensional Structure ◽  
Directed Mutagenesis ◽  
Catalytic Residue ◽  
Amino Acid Residues ◽  
Insight Into

ABSTRACT A novel FAD-dependent thymidylate synthase, ThyX, is present in a variety of eubacteria and archaea, including the mycobacteria. A short motif found in all thyX genes, RHRX7-8S, has been identified. The three-dimensional structure of the Mycobacterium tuberculosis ThyX enzyme has been solved. Building upon this information, we used directed mutagenesis to produce 67 mutants of the M. tuberculosis thyX gene. Each enzyme was assayed to determine its ability to complement the defect in thymidine biosynthesis in a ΔthyA strain of Escherichia coli. Enzymes from selected strains were then tested in vitro for their ability to catalyze the oxidation of NADPH and the release of a proton from position 5 of the pyrimidine ring of dUMP. The results defined an extended motif of amino acids essential to enzyme activity in M. tuberculosis (Y44X24 H69X25R95HRX7 S105XRYX90R199 [with the underlined histidine acting as the catalytic residue and the underlined serine as the nucleophile]) and provided insight into the ThyX reaction mechanism. ThyX is found in a variety of bacterial pathogens but is absent in humans, which depend upon an unrelated thymidylate synthase, ThyA. Therefore, ThyX is a potential target for development of antibacterial drugs.

scholarly journals Structure Characteristics, Biochemical Properties, and Pharmaceutical Applications of Alginate Lyases

Marine Drugs ◽  
2021 ◽  
Vol 19 (11) ◽  
pp. 628
Author(s):  
Shu-Kun Gao ◽  
Rui Yin ◽  
Xiao-Chen Wang ◽  
Hui-Ning Jiang ◽  
Xiao-Xiao Liu ◽  
...  
Keyword(s):  
Brown Algae ◽  
Catalytic Mechanism ◽  
Degradation Products ◽  
Structural Characteristics ◽  
Catalytic Efficiency ◽  
Alginate Lyase ◽  
Biochemical Properties ◽  
Structure And Property ◽  
Alginate Oligosaccharides ◽  
Alginate Lyases

Alginate, the most abundant polysaccharides of brown algae, consists of various proportions of uronic acid epimers α-L-guluronic acid (G) and β-D-mannuronic acid (M). Alginate oligosaccharides (AOs), the degradation products of alginates, exhibit excellent bioactivities and a great potential for broad applications in pharmaceutical fields. Alginate lyases can degrade alginate to functional AOs with unsaturated bonds or monosaccharides, which can facilitate the biorefinery of brown algae. On account of the increasing applications of AOs and biorefinery of brown algae, there is a scientific need to explore the important aspects of alginate lyase, such as catalytic mechanism, structure, and property. This review covers fundamental aspects and recent developments in basic information, structural characteristics, the structure–substrate specificity or catalytic efficiency relationship, property, molecular modification, and applications. To meet the needs of biorefinery systems of a broad array of biochemical products, alginate lyases with special properties, such as salt-activated, wide pH adaptation range, and cold adaptation are outlined. Withal, various challenges in alginate lyase research are traced out, and future directions, specifically on the molecular biology part of alginate lyases, are delineated to further widen the horizon of these exceptional alginate lyases.

scholarly journals Conformation and membrane interaction studies of the potent antimicrobial and anticancer peptide palustrin-Ca

2021 ◽  
Author(s):  
Patrick Brendan Timmons ◽  
Chandralal M Hewage
Keyword(s):  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Dynamics Simulation ◽  
Peptide Sequence ◽  
Helical Conformation ◽  
Dimensional Structure ◽  
Amino Acid Residues ◽  
Anticancer Activities ◽  
American Bullfrog ◽  
Α Helix

Palustrin-Ca (GFLDIIKDTGKEFAVKILNNLKCKLAGGCPP) is a host defense peptide with potent antimicrobial and anticancer activities, first isolated from the skin of the American bullfrog Lithobates catesbeianus. The peptide is 31 amino acid residues long, cationic and amphipathic. Two-dimensional NMR spectroscopy was employed to characterise its three-dimensional structure in a 50/50% water/2,2,2-trifluoroethanol-d3 mixture. The structure is defined by an α-helix that spans between Ile6-Ala26, and a cyclic disulphide bridged domain at the C-terminal end of the peptide sequence, between residues 23 and 29. A molecular dynamics simulation was employed to model the peptide's interactions with sodium dodecyl sulphate micelles, a widely used bacterial membrane-mimicking environment. Throughout the simulation, the peptide was found to maintain its α-helical conformation between residues Ile6-Ala26, while adopting a position parallel to the surface to micelle, which is energetically-favourable due to many hydrophobic and electrostatic contacts with the micelle.

scholarly journals Expression and Characterization of an Alginate Lyase and Its Thermostable Mutant in Pichia pastoris

Marine Drugs ◽  
2020 ◽  
Vol 18 (6) ◽  
pp. 305
Author(s):  
Suxiao Yang ◽  
Zhemin Liu ◽  
Xiaodan Fu ◽  
Changliang Zhu ◽  
Qing Kong ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Degradation Products ◽  
Alginate Lyase ◽  
Maximal Activity ◽  
Degree Of Polymerization ◽  
Molecular Weights ◽  
Industrial Preparation ◽  
Alginate Oligosaccharides ◽  
Elimination Mechanism ◽  
G Ratio

Alginate is one of the most abundant polysaccharides in algae. Alginate lyase degrades alginate through a β-elimination mechanism to produce alginate oligosaccharides with special bioactivities. Improving enzyme activity and thermal stability can promote the application of alginate lyase in the industrial preparation of alginate oligosaccharides. In this study, the recombinant alginate lyase cAlyM and its thermostable mutant 102C300C were expressed and characterized in Pichia pastoris. The specific activities of cAlyM and 102C300C were 277.1 U/mg and 249.6 U/mg, respectively. Both enzymes showed maximal activity at 50 °C and pH 8.0 and polyG preference. The half-life values of 102C300C at 45 °C and 50 °C were 2.6 times and 11.7 times the values of cAlyM, respectively. The degradation products of 102C300C with a lower degree of polymerization contained more guluronate. The oligosaccharides with a polymerization degree of 2–4 were the final hydrolytic products. Therefore, 102C300C is potentially valuable in the production of alginate oligosaccharides with specific M/G ratio and molecular weights.

Characteristics of Two Forms of α-Amylases and Structural Implication

1999 ◽  
Vol 65 (10) ◽  
pp. 4652-4658 ◽  
Author(s):  
Kohji Ohdan ◽  
Takashi Kuriki ◽  
Hiroki Kaneko ◽  
Jiro Shimada ◽  
Toshikazu Takada ◽  
...  
Keyword(s):  
Amino Acid ◽  
Three Dimensional ◽  
Amino Acid Sequences ◽  
Dimensional Structure ◽  
Amino Acid Residues ◽  
Amylase Gene ◽  
Raw Starch ◽  
Raw Starch Binding ◽  
Terminal Amino ◽  
Carboxyl Terminal

ABSTRACT Complete (Ba-L) and truncated (Ba-S) forms of α-amylases fromBacillus subtilis X-23 were purified, and the amino- and carboxyl-terminal amino acid sequences of Ba-L and Ba-S were determined. The amino acid sequence deduced from the nucleotide sequence of the α-amylase gene indicated that Ba-S was produced from Ba-L by truncation of the 186 amino acid residues at the carboxyl-terminal region. The results of genomic Southern analysis and Western analysis suggested that the two enzymes originated from the same α-amylase gene and that truncation of Ba-L to Ba-S occurred during the cultivation of B. subtilis X-23 cells. Although the primary structure of Ba-S was approximately 28% shorter than that of Ba-L, the two enzyme forms had the same enzymatic characteristics (molar catalytic activity, amylolytic pattern, transglycosylation ability, effect of pH on stability and activity, optimum temperature, and raw starch-binding ability), except that the thermal stability of Ba-S was higher than that of Ba-L. An analysis of the secondary structure as well as the predicted three-dimensional structure of Ba-S showed that Ba-S retained all of the necessary domains (domains A, B, and C) which were most likely to be required for functionality as α-amylase.

scholarly journals Efficacy of Dideoxynucleosides against Human Foamy Virus and Relationship to Its Reverse Transcriptase Amino Acid Sequence and Structure

2001 ◽  
Vol 75 (15) ◽  
pp. 7184-7187 ◽  
Author(s):  
Anne Yvon-Groussin ◽  
Pierre Mugnier ◽  
Philippe Bertin ◽  
Marc Grandadam ◽  
Henri Agut ◽  
...  
Keyword(s):  
Amino Acid ◽  
Reverse Transcriptase ◽  
Foamy Virus ◽  
Three Dimensional ◽  
Retroviral Vectors ◽  
Dimensional Structure ◽  
Human Foamy Virus ◽  
Amino Acid Residues ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Human foamy virus (HFV), a retrovirus of simian origin which occasionally infects humans, is the basis of retroviral vectors in development for gene therapy. Clinical considerations of how to treat patients developing an uncontrolled infection by either HFV or HFV-based vectors need to be raised. We determined the susceptibility of the HFV to dideoxynucleosides and found that only zidovudine was equally efficient against the replication of human immunodeficiency virus type 1 (HIV-1) and HFV. By contrast, zalcitabine (ddC), lamivudine (3TC), stavudine (d4T), and didanosine (ddI) were 3-, 3-, 30-, and 46-fold less efficient against HFV than against HIV-1, respectively. Some amino acid residues known to be involved in HIV-1 resistance to ddC, 3TC, d4T, and ddI were found at homologous positions of HFV reverse transcriptase (RT). These critical amino acids are located at the same positions in the three-dimensional structure of HIV-1 and HFV RT, suggesting that both enzymes share common patterns of inhibition.

scholarly journals Secretory Expression of an Alkaline Alginate Lyase With Heat Recovery Property in Yarrowia lipolytica

2021 ◽  
Vol 12 ◽  
Author(s):  
Lu Liu ◽  
Zhipeng Wang ◽  
Zhihong Zheng ◽  
Ze Li ◽  
Xiaofeng Ji ◽  
...  
Keyword(s):  
Amino Acids ◽  
Yarrowia Lipolytica ◽  
Heat Recovery ◽  
Degradation Products ◽  
Biological Activities ◽  
Alginate Lyase ◽  
Potential Candidate ◽  
Product Specificity ◽  
Alginate Oligosaccharides ◽  
Recovery Performance

Alginate lyase possesses wide application prospects for the degradation of brown algae and preparation of alginate oligosaccharides, and its degradation products display a variety of biological activities. Although many enzymes of this type have been reported, alginate lyases with unique properties are still relatively rare. In the present work, an alginate lyase abbreviated as Alyw203 has been cloned from Vibrio sp. W2 and expressed in food-grade Yarrowia lipolytica. The Alyw203 gene consists of an open reading frame (ORF) of 1,566 bp containing 521 amino acids, of which the first 17 amino acids are considered signal peptides, corresponding to secretory features. The peak activity of the current enzyme appears at 45°C with a molecular weight of approximately 57.0 kDa. Interestingly, Alyw203 exhibits unique heat recovery performance, returning above 90% of its initial activity in the subsequent incubation for 20 min at 10°C, which is conducive to the recovery of current enzymes at low-temperature conditions. Meanwhile, the highest activity is obtained under alkaline conditions of pH 10.0, showing outstanding pH stability. Additionally, as an alginate lyase independent of NaCl and resistant to metal ions, Alyw203 is highly active in various ionic environments. Moreover, the hydrolyzates of present enzymes are mainly concentrated in the oligosaccharides of DP1–DP2, displaying perfect product specificity. The alkali suitability, heat recovery performance, and high oligosaccharide yield of Alyw203 make it a potential candidate for industrial production of the monosaccharide and disaccharide.

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