scholarly journals Thermostable D-amino acid decarboxylases derived from Thermotoga maritima diaminopimelate decarboxylase

2021 ◽  
Vol 34 ◽  
Author(s):  
Antonija Marjanovic ◽  
Carlos J Ramírez-Palacios ◽  
Marcelo F Masman ◽  
Jeroen Drenth ◽  
Marleen Otzen ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Building Block ◽  
Catalytic Properties ◽  
Thermotoga Maritima ◽  
Aminocaproic Acid ◽  
Biosynthetic Pathways ◽  
Catalytic Activities ◽  
The Common ◽  
Diaminopimelate Decarboxylase

Abstract Diaminopimelate decarboxylases (DAPDCs) are highly selective enzymes that catalyze the common final step in different lysine biosynthetic pathways, i.e. the conversion of meso-diaminopimelate (DAP) to L-lysine. We examined the modification of the substrate specificity of the thermostable decarboxylase from Thermotoga maritima with the aim to introduce activity with 2-aminopimelic acid (2-APA) since its decarboxylation leads to 6-aminocaproic acid (6-ACA), a building block for the synthesis of nylon-6. Structure-based mutagenesis of the distal carboxylate binding site resulted in a set of enzyme variants with new activities toward different D-amino acids. One of the mutants (E315T) had lost most of its activity toward DAP and primarily acted as a 2-APA decarboxylase. We next used computational modeling to explain the observed shift in catalytic activities of the mutants. The results suggest that predictive computational protocols can support the redesign of the catalytic properties of this class of decarboxylating PLP-dependent enzymes.

scholarly journals A Practical Approach to the Investigation of Amino Acid Disorders

1988 ◽  
Vol 25 (2) ◽  
pp. 129-141 ◽  
Author(s):  
M A Edwards ◽  
S Grant ◽  
A Green
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Analysis ◽  
Practical Approach ◽  
The Common ◽  
Common Problems

We have, in this paper, highlighted some of the common problems in amino acid analysis in our experience and listed the possible causes for increases in specific amino acids in urine—together with guidance on appropriate follow-up investigations.

Renal reabsorption of amino acids

1962 ◽  
Vol 203 (5) ◽  
pp. 891-896 ◽  
Author(s):  
Marian Ruszkowski ◽  
Cizesław Arasimowicz ◽  
Jan Knapowski ◽  
Jan Steffen ◽  
Krystyna Weiss
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Transport Mechanism ◽  
Flow Analysis ◽  
Proximal Segment ◽  
Renal Tubules ◽  
Basic Amino Acids ◽  
Tubular Transport ◽  
The Common ◽  
Stop Flow

Using the method of stop flow analysis an attempt was made to localize the process of amino acid reabsorption in the nephron of the dog. Special attention was given to the group of basic amino acids and cystine believed to share a common tubular transport mechanism. The evidence obtained in this study points clearly to the proximal segment as the site of intensive reabsorption of all amino acids investigated. During the infusion of arginine, lysine or ornithine, an increased excretion of two remaining basic amino acids plus cystine was observed, as a rule. Successful attempts were made to infuse cystine intravenously. The results of these experiments did provide the missing link for the hypothesis derived by Dent and Rose ( Quart. J. Med. 20: 205, 1951) concerning the common transport mechanism of arginine, ornithine, lysine, and cystine in the renal tubules. The functional cystinuria, which can be induced by saturating the common reabsorptive pathway with each of the above-mentioned amino acids, is fully reversible.

scholarly journals Facilitating the Evolution of Esterase Activity from a Promiscuous Enzyme (Mhg) with Catalytic Functions of Amide Hydrolysis and Carboxylic Acid Perhydrolysis by Engineering the Substrate Entrance Tunnel

2016 ◽  
Vol 82 (22) ◽  
pp. 6748-6756 ◽  
Author(s):  
Xiaodan Yan ◽  
Jianjun Wang ◽  
Yu Sun ◽  
Junge Zhu ◽  
Sheng Wu
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Engineering ◽  
Esterase Activity ◽  
Molecular Basis ◽  
Catalytic Triad ◽  
Amino Acid Substitutions ◽  
Biological Functions ◽  
Catalytic Activities ◽  
Efficient Strategy

ABSTRACTPromiscuous enzymes are generally considered to be starting points in the evolution of offspring enzymes with more specific or even novel catalytic activities, which is the molecular basis of producing new biological functions. Mhg, a typical α/β fold hydrolase, was previously reported to have both γ-lactamase and perhydrolase activities. However, despite having high structural similarity to and sharing an identical catalytic triad with an extensively studied esterase fromPseudomonas fluorescens, this enzyme did not show any esterase activity. Molecular docking and sequence analysis suggested a possible role for the entry of the binding pocket in blocking the entrance tunnel, preventing the ester compounds from entering into the pocket. By engineering the entrance tunnel with only one or two amino acid substitutions, we successfully obtained five esterase variants of Mhg. The variants exhibited a very broad substrate acceptance, hydrolyzing not only the classicalp-nitrophenol esters but also various types of chiral esters, which are widely used as drug intermediates. Site 233 at the entrance tunnel of Mhg was found to play a pivotal role in modulating the three catalytic activities by adjusting the size and shape of the tunnel, with different amino acid substitutions at this site facilitating different activities. Remarkably, the variant with the L233G mutation was a very specific esterase without any γ-lactamase and perhydrolase activities. Considering the amino acid conservation and differentiation, this site could be a key target for future protein engineering. In addition, we demonstrate that engineering the entrance tunnel is an efficient strategy to regulate enzyme catalytic capabilities.IMPORTANCEPromiscuous enzymes can act as starting points in the evolution of novel catalytic activities, thus providing a molecular basis for the production of new biological functions. In this study, we identified a critical amino acid residue (Leu233) at the entry of the substrate tunnel of a promiscuous enzyme, Mhg. We found that substitution of this residue with smaller amino acids such as Gly, Ala, Ser, or Pro endowed the enzyme with novel esterase activity. Different amino acids at this site can facilitate different catalytic activities. These findings exhibited universal significance in this subset of α/β fold hydrolases, including Mhg. Furthermore, we demonstrate that engineering the entrance tunnel is an efficient strategy to evolve new enzyme catalytic capabilities. Our study has important implications for the regulation of enzyme catalytic promiscuity and development of protein engineering methodologies.

scholarly journals The Rare Amino Acid Building Block 3-(3-furyl)-Alanine in the Formation of Non-ribosomal Peptides

2017 ◽  
Vol 12 (1) ◽  
pp. 1934578X1701200 ◽  
Author(s):  
Fayrouz El Maddah ◽  
Mamona Nazir ◽  
Gabriele M. König
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Secondary Metabolites ◽  
Cyclosporine A ◽  
Building Block ◽  
Therapeutic Potential ◽  
Rhizopus Microsporus ◽  
Bacterial Endosymbiont ◽  
Peptide Synthetases ◽  
Microbial Origin

Microorganisms have made considerable contributions to the production of peptide secondary metabolites, many of them with therapeutic potential, e.g., the fungus-derived immunosuppressant cyclosporine A and the antibiotic daptomycin originating from Streptomyces. Most of the medically used peptides are the product of non-ribosomal peptide synthetases (NRPS), incorporating apart from proteinogenic also unique, non-proteinogenic amino acids into the peptides. An extremely rare such amino acid is 3-(3-furyl)-alanine. So far, only few peptides have been found that contain this residue, including the rhizonins, bingchamide B and endolides. The producer of the rhizonins was proven to be the bacterial endosymbiont Burkholderia endofungorum inside the fungus Rhizopus microsporus. The microbial origin, chemistry and bioactivity of the 3-(3-furyl)-alanine containing peptides are the focus of this review.

Human 20S proteasome activity towards fluorogenic peptides of various chain lengths

2016 ◽  
Vol 397 (9) ◽  
pp. 921-926 ◽  
Author(s):  
Wioletta Rut ◽  
Marcin Drag
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Kinetic Analysis ◽  
20S Proteasome ◽  
Fluorogenic Substrates ◽  
Catalytic Activities ◽  
Peptide Length ◽  
Chain Lengths ◽  
Acid Substrate ◽  
Amino Acid Substrate

Abstract The proteasome is a multicatalytic protease responsible for the degradation of misfolded proteins. We have synthesized fluorogenic substrates in which the peptide chain was systematically elongated from two to six amino acids and evaluated the effect of peptide length on all three catalytic activities of human 20S proteasome. In the cases of five- and six-membered peptides, we have also synthesized libraries of fluorogenic substrates. Kinetic analysis revealed that six-amino-acid substrates are significantly better for chymotrypsin-like and caspase-like activity than shorter peptidic substrates. In the case of trypsin-like activity, a five-amino-acid substrate was optimal.

Development of 2-aminoisobutyric acid (Aib)-rich cell-penetrating foldamers for efficient siRNA delivery

2019 ◽  
Vol 55 (54) ◽  
pp. 7792-7795 ◽  
Author(s):  
Takashi Misawa ◽  
Nobumichi Ohoka ◽  
Makoto Oba ◽  
Hiroko Yamashita ◽  
Masakazu Tanaka ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Sirna Delivery ◽  
Aminoisobutyric Acid ◽  
Cell Penetrating ◽  
The Common

We have designed and synthesized a set of cell-penetrating foldamers (CPFs), Blocks 1–8, composed of the common amino acids Leu, Arg, and Gly, as well as the helicogenic amino acid 2-aminoisobutyric acid.

Cystathionine β-lyase is involved in d-amino acid metabolism

2018 ◽  
Vol 475 (8) ◽  
pp. 1397-1410 ◽  
Author(s):  
Tetsuya Miyamoto ◽  
Masumi Katane ◽  
Yasuaki Saitoh ◽  
Masae Sekine ◽  
Hiroshi Homma
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Acid Metabolism ◽  
Environmental Changes ◽  
Biosynthetic Pathways ◽  
E Coli ◽  
Comparable Level ◽  
Multifunctional Enzymes ◽  
Dehydratase Activity ◽  
Amino Acid Racemase

Non-canonical d-amino acids play important roles in bacteria including control of peptidoglycan metabolism and biofilm disassembly. Bacteria appear to produce non-canonical d-amino acids to adapt to various environmental changes, and understanding the biosynthetic pathways is important. We identified novel amino acid racemases possessing the ability to produce non-canonical d-amino acids in Escherichia coli and Bacillus subtilis in our previous study, whereas the biosynthetic pathways of these d-amino acids still remain unclear. In the present study, we demonstrated that two cystathionine β-lyases (MetC and MalY) from E. coli produce non-canonical d-amino acids including non-proteinogenic amino acids. Furthermore, MetC displayed d- and l-serine (Ser) dehydratase activity. We characterised amino acid racemase, Ser dehydratase and cysteine lyase activities, and all were higher for MetC. Interestingly, all three activities were at a comparable level for MetC, although optimal conditions for each reaction were distinct. These results indicate that MetC and MalY are multifunctional enzymes involved in l-methionine metabolism and the production of d-amino acids, as well as d- and l-Ser metabolism. To our knowledge, this is the first evidence that cystathionine β-lyase is a multifunctional enzyme with three different activities.

scholarly journals Retention and Loss of Amino Acid Biosynthetic Pathways Based on Analysis of Whole-Genome Sequences

2006 ◽  
Vol 5 (2) ◽  
pp. 272-276 ◽  
Author(s):  
Samuel H. Payne ◽  
William F. Loomis
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Minimal Medium ◽  
Whole Genome ◽  
Biosynthetic Pathways ◽  
Genome Sequences ◽  
Whole Genomes ◽  
Complete Genomes ◽  
Computational Analyses

ABSTRACT Plants and fungi can synthesize each of the 20 amino acids by using biosynthetic pathways inherited from their bacterial ancestors. However, the ability to synthesize nine amino acids (Phe, Trp, Ile, Leu, Val, Lys, His, Thr, and Met) was lost in a wide variety of eukaryotes that evolved the ability to feed on other organisms. Since the biosynthetic pathways and their respective enzymes are well characterized, orthologs can be recognized in whole genomes to understand when in evolution pathways were lost. The pattern of pathway loss and retention was analyzed in the complete genomes of three early-diverging protist parasites, the amoeba Dictyostelium, and six animals. The nine pathways were lost independently in animals, Dictyostelium, Leishmania, Plasmodium, and Cryptosporidium. Seven additional pathways appear to have been lost in one or another parasite, demonstrating that they are dispensable in a nutrition-rich environment. Our predictions of pathways retained and pathways lost based on computational analyses of whole genomes are validated by minimal-medium studies with mammals, fish, worms, and Dictyostelium. The apparent selective advantages of retaining biosynthetic capabilities for amino acids available in the diet are considered.

scholarly journals Phenotypic and Genotypic Analysis of Amino Acid Auxotrophy in Lactobacillus helveticus CNRZ 32

2007 ◽  
Vol 74 (2) ◽  
pp. 416-423 ◽  
Author(s):  
Jason K. Christiansen ◽  
Joanne E. Hughes ◽  
Dennis L. Welker ◽  
Beatriz T. Rodríguez ◽  
James L. Steele ◽  
...  
Keyword(s):  
Amino Acids ◽  
Lactic Acid ◽  
Amino Acid ◽  
Lactic Acid Bacteria ◽  
Ornithine Decarboxylase ◽  
Lactobacillus Helveticus ◽  
Decarboxylase Activity ◽  
Biosynthetic Pathways ◽  
Ornithine Decarboxylase Activity ◽  
Genotypic Analysis

ABSTRACT The conversion of amino acids into volatile and nonvolatile compounds by lactic acid bacteria in cheese is thought to represent the rate-limiting step in the development of mature flavor and aroma. Because amino acid breakdown by microbes often entails the reversible action of enzymes involved in biosynthetic pathways, our group investigated the genetics of amino acid biosynthesis in Lactobacillus helveticus CNRZ 32, a commercial cheese flavor adjunct that reduces bitterness and intensifies flavor notes. Most lactic acid bacteria are auxotrophic for several amino acids, and L. helveticus CNRZ 32 requires 14 amino acids. The reconstruction of amino acid biosynthetic pathways from a draft-quality genome sequence for L. helveticus CNRZ 32 revealed that amino acid auxotrophy in this species was due primarily to gene absence rather than point mutations, insertions, or small deletions, with good agreement between gene content and phenotypic amino acid requirements. One exception involved the phenotypic requirement for Asp (or Asn), which genome predictions suggested could be alleviated by citrate catabolism. This prediction was confirmed by the growth of L. helveticus CNRZ 32 after the addition of citrate to a chemically defined medium that lacked Asp and Asn. Genome analysis also predicted that L. helveticus CNRZ 32 possessed ornithine decarboxylase activity and would therefore catalyze the conversion of ornithine to putrescine, a volatile biogenic amine. However, experiments to confirm ornithine decarboxylase activity in L. helveticus CNRZ 32 by the use of several methods were unsuccessful, which indicated that this bacterium likely does not contribute to putrescine production in cheese.

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