DOCSLIB.ORG

Site-Specific Incorporation of Selenocysteine by Genetic Encoding As a Photocaged Unnatural Amino Acid Adarshi Welegedara, Luke A

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Site-Specific Incorporation of Selenocysteine by Genetic Encoding As a Photocaged Unnatural Amino Acid Adarshi Welegedara, Luke A Subscriber access provided by AUSTRALIAN NATIONAL UNIV Article Site-specific incorporation of selenocysteine by genetic encoding as a photocaged unnatural amino acid Adarshi Welegedara, Luke A. Adams, Bim Graham, Thomas Huber, and Gottfried Otting Bioconjugate Chem., Just Accepted Manuscript • DOI: 10.1021/acs.bioconjchem.8b00254 • Publication Date (Web): 06 Jun 2018 Downloaded from http://pubs.acs.org on June 12, 2018 Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts. is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties. Page 1 of 21 Bioconjugate Chemistry Site-specific incorporation of selenocysteine by genetic encoding as a photocaged unnatural amino 1 2 acid 3 4 5 1 2 1 2 1* 6 Adarshi P. Welegedara, Luke A. Adams, Thomas Huber, Bim Graham, Gottfried Otting 7 8 1 9 Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia 10 2 11 Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC 3052, Australia 12 13 14 *corresponding author 15 16 phone: +61 2 61256507 17 fax: +61 2 61250750 18 19 e-mail: [email protected] 20 21 22 23 ABSTRACT 24 Selenocysteine (Sec) is a naturally occurring amino acid that is also referred to as the 21st amino acid. 25 26 Site-specific incorporation of Sec into proteins is attractive, because the reactivity of a selenol group 27 28 exceeds that of a thiol group and thus allows site-specific protein modifications. It is incorporated into 29 30 proteins by an unusual enzymatic mechanism which, in E. coli and other organisms, involves the 31 recognition of a selenocysteine insertion sequence (SECIS) in the mRNA of the target protein. 32 33 Reengineering of the natural machinery for Sec incorporation at arbitrary sites independent of SECIS 34 35 elements, however, is challenging. Here we demonstrate an alternative route, whereby a photocaged 36 selenocysteine (PSc) is incorporated as an unnatural amino acid in response to an amber stop codon, 37 38 using a mutant Methanosarcina mazei pyrrolysyl-tRNA synthetase, Mm PCC2RS, and its cognate 39 40 tRNACUA. Following decaging by UV irradiation, proteins synthesized with PSc are readily tagged, e.g. 41 42 with NMR probes to study ligand binding by NMR spectroscopy. The approach provides a facile route 43 for genetically encoded Sec incorporation. It allows the production of pure selenoproteins and the Sec 44 45 residue enables site-specific covalent protein modification with reagents that would usually react first 46 47 with naturally occurring cysteine residues. The much greater reactivity of Sec residues allows their 48 selective alkylation in the presence of highly solvent-exposed cysteine residues. 49 50 51 52 53 54 55 56 57 58 59 60 ACS Paragon Plus Environment 1 Bioconjugate Chemistry Page 2 of 21 INTRODUCTION 1 2 3 Site-specific protein modification is a fundamental tool for a myriad of applications in protein 4 5 chemistry, including protein immobilization on surfaces, enhancement of protein solubility by 6 PEGylation, production of therapeutic conjugates with drugs, mimicking of natural post-translational 7 8 modifications, and introduction of tags for structural studies by fluorescence resonance energy transfer 9 10 (FRET), electron paramagnetic resonance (EPR) spectroscopy, or nuclear magnetic resonance (NMR) 11 spectroscopy.1−8 Cysteine is the most convenient target for protein modification as it is the most 12 13 nucleophilic and oxidation-sensitive among the 20 canonical amino acids. Thus site-directed 14 15 modifications of proteins are often carried out on cysteine residues with the formation of disulfide or 16 17 thioether bonds. To be site-selective, however, this approach requires that only a single cysteine residue 18 is amenable to the chemical reaction, which is difficult when the protein of interest contains other 19 20 cysteine residues that are essential for the structure or function of the protein. In principle, this problem 21 9 22 can be solved by genetic encoding of photocaged cysteine (PCC) as an unnatural amino acid (UAA), 23 24 which enables a protocol involving the chemical protection of the native cysteine residues, decaging of 25 the PCC residue and chemical modification and deprotection of the native cysteine residues.10 Charging 26 27 of the cognate amber suppressor tRNACUA with the PCC amino acid is elegantly achieved by a mutant 28 9 29 Methanosarcina barkeri pyrrolysyl-tRNA synthetase (Mb PylRS), named PCC2RS, but the number of 30 steps involved in selective cysteine labeling renders the overall process unattractive. A more 31 32 straightforward alternative is to incorporate a Sec residue, which is chemically more reactive than 33 34 cysteine, obviating the need for protection of any native cysteine residues. Sec is chemically similar to 35 11−13 36 cysteine but has a significantly lower pKa value (5.2 versus 8.5). This can be exploited for selective 37 modifications of Sec residues in the presence of native cysteine residues at slightly acidic pH values.14- 38 39 −17 A recent, very attractive application harnesses the exceptional reactivity of Sec residues for 40 17−19 41 expressed protein-ligation. 42 43 Site-specific incorporation of Sec is challenging. The natural mechanism employs a highly 44 specialized translation system, relying on a specific tRNA (tRNASec), selenophosphate synthase (SelD), 45 46 Sec synthase (SelA), a special EF-Tu-independent elongation factor (SelB), and a conserved mRNA 47 48 sequence (selenocysteine insertion sequence or SECIS) in the vicinity of the target UGA codon. The 49 requisite mRNA stem-loop structure comprised by the SECIS element restricts the sequence context 50 51 where Sec residues can be positioned.11,13 In an E. coli strain devoid of the release factor 1 (RF1), the 52 53 SECIS/SelB system has also successfully been harnessed for the synthesis of pure selenoproteins using 54 55 a UAG (amber) codon instead of the usual UGA codon but, in the absence of the SECIS element, this 56 approach results in full-length protein due to misincorporation of natural amino acids.20 Site-selective 57 58 Sec incorporation at an amber stop codon independent of SelB and the SECIS element has also been 59 60 ACS Paragon Plus Environment 2 Page 3 of 21 Bioconjugate Chemistry demonstrated, using a tRNASec engineered for EF-Tu–mediated Sec incorporation.21−24 At present, this 1 2 approach is limited to low yields of selenoproteins (<0.5 mg/L cell culture), and further optimization of 3 Sec 25 4 the engineered tRNA is difficult to conceive as it needs to be recognized both by SelA and EF-Tu. 5 Poor expression yields thus have prevented the use of this approach for subsequent applications that 6 7 require larger protein quantities, such as NMR spectroscopy or expressed protein ligation. The problem 8 9 of low yield is circumvented by a new system for EF-Tu mediated selenocysteine incorporation at 10 11 amber codons, which uses Aeromonas salmonicida selenocysteine synthase (SelA) to convert Ser-allo- 12 tRNA into Sec-allo-tRNA which is efficiently recognized by EF-Tu.26 The system can be used to obtain 13 14 selenoproteins in high yields, but it is prone to incorporation of serine in place of selenocysteine due to 15 16 incomplete conversion of Ser to Sec by SelA. This makes the strategy critically dependent on 17 18 optimization of the relative expression levels of allo-tRNA and SelA as well as the expression 26 19 temperatures, which are different for different proteins. 20 21 The wide interest in Sec incorporation has triggered the development of a range of alternative 22 23 strategies for the production of selenoproteins, in particular focusing on expression in E. coli, which is 24 the most readily used organism for protein production. In a straightforward way, cysteine residues can 25 26 globally be replaced by Sec by misloading the cysteinyl-tRNA with Sec in a growth medium 27 28 supplemented with Sec, but incorporation of cysteine cannot be completely avoided in this approach and 29 27 30 site-selectivity is achieved only if the native protein sequence contains only a single cysteine residue, 31 or a Sec-containing fragment is post-translationally ligated with a separately produced polypeptide.17 As 32 33 an alternative, it has been shown that a photocaged Sec, 4,5-dimethoxy-2-nitrobenzyl-Sec (DMNB-Sec), 34 35 can be incorporated as a UAA in yeast by genetic encoding based on an orthogonal LeuRS/tRNACUA 36 pair, with subsequent decaging providing a straightforward route to a site-specific Sec residue.16 This 37 38 system, however, requires high concentrations of DMNB-Sec (3 mM) to suppress erroneous 39 16 40 incorporation of leucine and isoleucine, and cannot be used in E.
Load more

Recommended publications
  • Selenocysteine, Pyrrolysine, and the Unique Energy Metabolism of Methanogenic Archaea
    Hindawi Publishing Corporation Archaea Volume 2010, Article ID 453642, 14 pages doi:10.1155/2010/453642 Review Article Selenocysteine, Pyrrolysine, and the Unique Energy Metabolism of Methanogenic Archaea Michael Rother1 and Joseph A. Krzycki2 1 Institut fur¨ Molekulare Biowissenschaften, Molekulare Mikrobiologie & Bioenergetik, Johann Wolfgang Goethe-Universitat,¨ Max-von-Laue-Str. 9, 60438 Frankfurt am Main, Germany 2 Department of Microbiology, The Ohio State University, 376 Biological Sciences Building 484 West 12th Avenue Columbus, OH 43210-1292, USA Correspondence should be addressed to Michael Rother, [email protected] andJosephA.Krzycki,[email protected] Received 15 June 2010; Accepted 13 July 2010 Academic Editor: Jerry Eichler Copyright © 2010 M. Rother and J. A. Krzycki. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Methanogenic archaea are a group of strictly anaerobic microorganisms characterized by their strict dependence on the process of methanogenesis for energy conservation. Among the archaea, they are also the only known group synthesizing proteins containing selenocysteine or pyrrolysine. All but one of the known archaeal pyrrolysine-containing and all but two of the confirmed archaeal selenocysteine-containing protein are involved in methanogenesis. Synthesis of these proteins proceeds through suppression of translational stop codons but otherwise the two systems are fundamentally different. This paper highlights these differences and summarizes the recent developments in selenocysteine- and pyrrolysine-related research on archaea and aims to put this knowledge into the context of their unique energy metabolism. 1. Introduction found to correspond to pyrrolysine in the crystal structure [9, 10] and have its own tRNA [11].
    [Show full text]
  • Selenocysteine, Identified As the Penultimate C-Terminal Residue in Human T-Cell Thioredoxin Reductase, Corresponds to TGA in the Human Placental Gene" (1996)
    University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln Vadim Gladyshev Publications Biochemistry, Department of June 1996 Selenocysteine, identified as the penultimate C-terminal esiduer in human T-cell thioredoxin reductase, corresponds to TGA in the human placental gene Vadim Gladyshev University of Nebraska-Lincoln, [email protected] Kuan-Teh Jeang National Institutes of Health, Bethesda, MD Thressa C. Stadtman National Institutes of Health, Bethesda, MD Follow this and additional works at: https://digitalcommons.unl.edu/biochemgladyshev Part of the Biochemistry, Biophysics, and Structural Biology Commons Gladyshev, Vadim; Jeang, Kuan-Teh; and Stadtman, Thressa C., "Selenocysteine, identified as the penultimate C-terminal residue in human T-cell thioredoxin reductase, corresponds to TGA in the human placental gene" (1996). Vadim Gladyshev Publications. 23. https://digitalcommons.unl.edu/biochemgladyshev/23 This Article is brought to you for free and open access by the Biochemistry, Department of at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in Vadim Gladyshev Publications by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. Proc. Natl. Acad. Sci. USA Vol. 93, 6146-6151, June 1996 Biochemistrypp. Selenocysteine, identified as the penultimate C-terminal residue in human T-cell thioredoxin reductase, corresponds to TGA in the human placental gene (selenium/thioredoxin reductase/TGA/selenocysteine) VADIM N. GLADYSHEV*, KUAN-TEH JEANGt, AND THRESSA C. STADTMAN*t *Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, and tLaboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 9000 Rockville Pike, Bethesda, MD 20892 Contributed by Thressa C. Stadtman, February 27, 1996 ABSTRACT The possible relationship of selenium to im- peroxidase family (8).
    [Show full text]
  • Characterization of a Selenocysteine-Ligated P450 Compound I Reveals Direct Link Between Electron Donation and Reactivity Elizab
    Characterization of a Selenocysteine-ligated P450 Compound I Reveals Direct Link Between Electron Donation and Reactivity Elizabeth Onderko†, Alexey Silakov†, Timothy H. Yosca‡, and Michael T. Green‡,* ‡Departments of Chemistry & Molecular Biology and Biochemistry, University of California, Irvine, CA 92697 †Department of Chemistry, Penn State University, University Park, PA 16802 Contact: [email protected] Abstract Strong electron-donation from the axial thiolate-ligand of cytochrome P450 has been proposed to increase the reactivity of compound I with respect to C–H bond activation. However, it has proven difficult to test this hypothesis, and a direct link between reactivity and electron donation has yet to be established. To make this connection, we have prepared a selenolate-ligated cytochrome P450 compound I intermediate. This isoelectronic perturbation allows for direct comparisons with the wild type enzyme. Selenium incorporation was obtained using a cysteine auxotrophic E. coli strain. The intermediate was prepared with meta-chloroperbenzoic acid and characterized by UV-visible, Mössbauer, and electron paramagnetic resonance spectroscopies. Measurements revealed increased asymmetry around the ferryl moiety, consistent with increased electron donation from the axial selenolate-ligand. In line with this observation, we find that the selenolate-ligated compound I cleaves C–H bonds more rapidly than the wild-type intermediate. Background Cytochrome P450s are a class of thiolate-ligated heme proteins that are known for their ability to functionalize unactivated C–H bonds. In efforts to understand reactivity in these and other thiolate-heme systems, comparisons are often drawn between P450s and the histidine-ligated heme peroxidases. Both classes of heme enzymes share a similar active intermediate: a ferryl (or iron(IV)oxo) radical species, called compound I1-3.
    [Show full text]
  • Generation of Recombinant Mammalian Selenoproteins Through Ge- Netic Code Expansion with Photocaged Selenocysteine
    bioRxiv preprint doi: https://doi.org/10.1101/759662; this version posted September 5, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Generation of Recombinant Mammalian Selenoproteins through Ge- netic Code Expansion with Photocaged Selenocysteine. Jennifer C. Peeler, Rachel E. Kelemen, Masahiro Abo, Laura C. Edinger, Jingjia Chen, Abhishek Chat- terjee*, Eranthie Weerapana* Department of Chemistry, Boston College, Chestnut Hill, Massachusetts 02467, United States Supporting Information Placeholder ABSTRACT: Selenoproteins contain the amino acid sele- neurons susceptible to ferroptotic cell death due to nocysteine and are found in all domains of life. The func- overoxidation and inactivation of GPX4-Cys.4 This ob- tions of many selenoproteins are poorly understood, servation demonstrates a potential advantage conferred partly due to difficulties in producing recombinant sele- by the energetically expensive production of selenopro- noproteins for cell-biological evaluation. Endogenous teins. mammalian selenoproteins are produced through a non- Sec incorporation deviates from canonical protein canonical translation mechanism requiring suppression of translation, requiring suppression of the UGA stop codon. the UGA stop codon, and a selenocysteine insertion se- In eukaryotes, Sec biosynthesis occurs directly on the quence (SECIS) element in the 3’ untranslated region of suppressor tRNA (tRNA[Ser]Sec). Specifically, tRNA[Ser]Sec the mRNA. Here, recombinant selenoproteins are gener- is aminoacylated with serine by seryl-tRNA synthetase ated in mammalian cells through genetic code expansion, (SerS), followed by phosphorylation by phosphoseryl- circumventing the requirement for the SECIS element, tRNA kinase (PSTK), and subsequent Se incorporation and selenium availability.
    [Show full text]
  • Site-Specific Protein Modifications Through Pyrroline-Carboxy-Lysine Residues
    Site-specific protein modifications through pyrroline-carboxy-lysine residues Weijia Ou1, Tetsuo Uno1, Hsien-Po Chiu, Jan Grünewald, Susan E. Cellitti, Tiffany Crossgrove, Xueshi Hao, Qian Fan, Lisa L. Quinn, Paula Patterson, Linda Okach, David H. Jones, Scott A. Lesley, Ansgar Brock, and Bernhard H. Geierstanger2 Genomics Institute of the Novartis Research Foundation, 10675 John-Jay-Hopkins Drive, San Diego, CA 92121-1125 Edited* by Peter G. Schultz, The Scripps Research Institute, La Jolla, CA, and approved May 11, 2011 (received for review April 4, 2011) Pyrroline-carboxy-lysine (Pcl) is a demethylated form of pyrrolysine acids will face similar hurdles to achieve site-specificity without that is generated by the pyrrolysine biosynthetic enzymes when deleterious effects to the protein of interest. the growth media is supplemented with D-ornithine. Pcl is readily The most elegant way to generate homogenously, site-specifi- incorporated by the unmodified pyrrolysyl-tRNA/tRNA synthetase cally modified proteins is the in vivo incorporation of unnatural pair into proteins expressed in Escherichia coli and in mammalian amino acids (7–9). Over 70 unnatural amino acids featuring a cells. Here, we describe a broadly applicable conjugation chemistry wide array of functionalities can be incorporated at TAG codons that is specific for Pcl and orthogonal to all other reactive groups using specific tRNA/tRNA synthetase pairs engineered through on proteins. The reaction of Pcl with 2-amino-benzaldehyde or an in vivo selection process to be orthogonal to the cellular 2-amino-acetophenone reagents proceeds to near completion at machinery of the host cells. A set of reactive unnatural amino neutral pH with high efficiency.
    [Show full text]
  • Selenium Vs. Sulfur: Investigating the Substrate Specificity of a Selenocysteine Lyase
    University of Central Florida STARS Electronic Theses and Dissertations, 2004-2019 2019 Selenium vs. Sulfur: Investigating the Substrate Specificity of a Selenocysteine Lyase Michael Johnstone University of Central Florida Part of the Biotechnology Commons Find similar works at: https://stars.library.ucf.edu/etd University of Central Florida Libraries http://library.ucf.edu This Masters Thesis (Open Access) is brought to you for free and open access by STARS. It has been accepted for inclusion in Electronic Theses and Dissertations, 2004-2019 by an authorized administrator of STARS. For more information, please contact [email protected]. STARS Citation Johnstone, Michael, "Selenium vs. Sulfur: Investigating the Substrate Specificity of a Selenocysteine Lyase" (2019). Electronic Theses and Dissertations, 2004-2019. 6511. https://stars.library.ucf.edu/etd/6511 SELENIUM VS. SULFUR: INVESTIGATING THE SUBSTRATE SPECIFICITY OF A SELENOCYSTEINE LYASE by MICHAEL ALAN JOHNSTONE B.S. University of Central Florida, 2017 A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science in the Burnett School of Biomedical Sciences in the College of Medicine at the University of Central Florida Orlando, Florida Summer Term 2019 Major Professor: William T. Self © 2019 Michael Alan Johnstone ii ABSTRACT Selenium is a vital micronutrient in many organisms. While traces are required for survival, excess amounts are toxic; thus, selenium can be regarded as a biological “double-edged sword”. Selenium is chemically similar to the essential element sulfur, but curiously, evolution has selected the former over the latter for a subset of oxidoreductases. Enzymes involved in sulfur metabolism are less discriminate in terms of preventing selenium incorporation; however, its specific incorporation into selenoproteins reveals a highly discriminate process that is not completely understood.
    [Show full text]
  • Virtual 2-D Map of the Fungal Proteome
    www.nature.com/scientificreports OPEN Virtual 2‑D map of the fungal proteome Tapan Kumar Mohanta1,6*, Awdhesh Kumar Mishra2,6, Adil Khan1, Abeer Hashem3,4, Elsayed Fathi Abd‑Allah5 & Ahmed Al‑Harrasi1* The molecular weight and isoelectric point (pI) of the proteins plays important role in the cell. Depending upon the shape, size, and charge, protein provides its functional role in diferent parts of the cell. Therefore, understanding to the knowledge of their molecular weight and charges is (pI) is very important. Therefore, we conducted a proteome‑wide analysis of protein sequences of 689 fungal species (7.15 million protein sequences) and construct a virtual 2‑D map of the fungal proteome. The analysis of the constructed map revealed the presence of a bimodal distribution of fungal proteomes. The molecular mass of individual fungal proteins ranged from 0.202 to 2546.166 kDa and the predicted isoelectric point (pI) ranged from 1.85 to 13.759 while average molecular weight of fungal proteome was 50.98 kDa. A non‑ribosomal peptide synthase (RFU80400.1) found in Trichoderma arundinaceum was identifed as the largest protein in the fungal kingdom. The collective fungal proteome is dominated by the presence of acidic rather than basic pI proteins and Leu is the most abundant amino acid while Cys is the least abundant amino acid. Aspergillus ustus encodes the highest percentage (76.62%) of acidic pI proteins while Nosema ceranae was found to encode the highest percentage (66.15%) of basic pI proteins. Selenocysteine and pyrrolysine amino acids were not found in any of the analysed fungal proteomes.
    [Show full text]
  • Rare, but Essential – the Amino Acid Selenocysteine
    Molecular Biology Rare, but essential – the amino acid selenocysteine Based at the University of Bonn, Germany, Professor Dr Ulrich Schweizer is leading research into revealing the role of selenoproteins in mammalian physiology. By elucidating the mechanisms underlying their function, his work is yielding new insights into a wide array of human diseases affecting the brain and thyroid hormones. At the core of Prof Dr Schweizer’s research is the rare selenium-containing 21st amino acid selenocysteine (Sec), the defining component of selenoproteins. elenocysteine (Sec) is an the trace element selenium (Se) replaces the essential amino acid component sulphur atom of cysteine. Selenium possesses in selenoproteins, which are similar but more reactive properties than involved in a variety of cellular and sulphur, and is always housed in the active metabolic processes. Increasingly, centre of selenoenzymes. Stheir deregulation is being associated with neurodegenerative and other diseases, INVESTIGATING THE RAREST AMINO for which the underlying mechanisms ACID have remained unclear. However, using Sec eluded the Nobel Prize winning scientists novel transgenic mouse models, Prof Dr who deciphered the genetic code, because Schweizer and his team have uncovered Sec is encoded by what has been regarded a wealth of information regarding the exclusively as a termination codon, UGA. mechanisms behind their function. His team’s How can the cell distinguish between extensive research has found that reducing termination and Sec incorporation? There is a the expression of selenoproteins in the specific element within the mRNA sequence mammalian brain impairs brain development that directs the re-coding of UGA, called the and healthy functioning, consequently selenocysteine insertion sequence (SECIS).
    [Show full text]
  • Modern Diversification of the Amino Acid Repertoire Driven by Oxygen
    Modern diversification of the amino acid repertoire driven by oxygen Matthias Granolda, Parvana Hajievab, Monica Ioana Tos¸ac, Florin-Dan Irimiec, and Bernd Moosmanna,1 aEvolutionary Biochemistry and Redox Medicine, Institute for Pathobiochemistry, University Medical Center of the Johannes Gutenberg University, 55128 Mainz, Germany; bCellular Adaptation Group, Institute for Pathobiochemistry, University Medical Center of the Johannes Gutenberg University, 55128 Mainz, Germany; and cGroup of Biocatalysis and Biotransformations, Faculty of Chemistry and Chemical Engineering, Babes¸-Bolyai University, Cluj-Napoca 400028, Romania Edited by Harry B. Gray, California Institute of Technology, Pasadena, CA, and approved November 21, 2017 (received for review October 1, 2017) All extant life employs the same 20 amino acids for protein bio- aminoacyl-tRNA synthetases may not have been present in synthesis. Studies on the number of amino acids necessary to LUCA, the last universal common ancestor, but rather would have produce a foldable and catalytically active polypeptide have shown been distributed throughout life at a later time by lateral gene that a basis set of 7–13 amino acids is sufficient to build major struc- transfer (14). Similarly, the capacity to distinguish the late AA tural elements of modern proteins. Hence, the reasons for the evo- methionine from its genetic code neighbor isoleucine has probably lutionary selection of the current 20 amino acids out of a much larger developed only after the radiation of life (15). These observations available pool have remained elusive. Here, we have analyzed the call for selective factors behind the addition of methionine, Y, and quantum chemistry of all proteinogenic and various prebiotic amino W to the genetic code that have only become relevant to life acids.
    [Show full text]
  • Selenocysteine in Proteins—Properties and Biotechnological Use
    Biochimica et Biophysica Acta 1726 (2005) 1 – 13 http://www.elsevier.com/locate/bba Review Selenocysteine in proteins—properties and biotechnological use Linda Johanssona, Guro Gafvelinb, Elias S.J. Arne´ra,* aMedical Nobel Institute for Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institute, SE-171 77 Stockholm, Sweden bDepartment of Medicine, Clinical Immunology and Allergy Unit, Karolinska Institute and University Hospital, Stockholm, Sweden Received 12 April 2005; received in revised form 4 May 2005; accepted 7 May 2005 Available online 1 June 2005 Abstract Selenocysteine (Sec), the 21st amino acid, exists naturally in all kingdoms of life as the defining entity of selenoproteins. Sec is a cysteine (Cys) residue analogue with a selenium-containing selenol group in place of the sulfur-containing thiol group in Cys. The selenium atom gives Sec quite different properties from Cys. The most obvious difference is the lower pKa of Sec, and Sec is also a stronger nucleophile than Cys. Proteins naturally containing Sec are often enzymes, employing the reactivity of the Sec residue during the catalytic cycle and therefore Sec is normally essential for their catalytic efficiencies. Other unique features of Sec, not shared by any of the other 20 common amino acids, derive from the atomic weight and chemical properties of selenium and the particular occurrence and properties of its stable and radioactive isotopes. Sec is, moreover, incorporated into proteins by an expansion of the genetic code as the translation of selenoproteins involves the decoding of a UGA codon, otherwise being a termination codon. In this review, we will describe the different unique properties of Sec and we will discuss the prerequisites for selenoprotein production as well as the possible use of Sec introduction into proteins for biotechnological applications.
    [Show full text]
  • Structure and Functions of Amino Acids and Proteins
    Structure of Amino Acids DR. KIRAN MEENA 05/9/2019 8:00-9:00 AM Specific Learning Objectives 1. General Structure of amino acids 2. Amino acids classification based on: •Standard and Non-standard amino acids (aa) •Essential and non-essential aa •Ketogenic and Glucogenic aa •Side chain functional group 3. Function of essential amino acids Introduction •Amino acids as a building blocks of peptides and proteins •Proteins are made up of hundreds of smaller units called amino acids that are attached to one another by peptide bonds, forming a long chain. •Protein as a string of beads where each bead is an amino acid. www.khanacademy.org Genetic Code Specifies 20 L-α-Amino Acids •Proteins are synthesized from the set of 20 L-α-amino acids encoded by nucleotide triplets called codons. •Common amino acids are those for which at least one specific codon exists in the DNA genetic code. •Sequences of peptides and proteins represent by using one- and three letter abbreviations for each amino acid. Genetic information is transcribed from a DNA sequence into mRNA and then translated to amino acid sequence of a protein Fig. 2.1. Textbook of Biochemistry with Clinical Correlations, 4th edition by Thomas M Devlin General Structure of Common Amino Acids •General structure of amino acids , group and a variable side chain •Side chain determines: protein folding, binding to specific ligand and interaction with its environment •Amino acids consists of a constant COOH (R is side chain) + - •At neutral pH, H2N- protonated to H3N -, and –COOH deprotonated to –COO Fig.4.2.
    [Show full text]
  • Towards the Characterization of the Eukaryotic Selenoproteome: A
    Towards the characterization of the eukaryotic selenoproteome a computational approach Sergi Castellano Hereza Ph.D. dissertation Barcelona, May 2004 Departament de Ciencies` Experimentals i de la Salut Universitat Pompeu Fabra Towards the characterization of the eukaryotic selenoproteome a computational approach Sergi Castellano Hereza Memoria` presentada per optar al grau de Doctor en Biologia per la Universitat Pompeu Fabra. Aquesta Tesi Doctoral ha estat realitzada sota la direccio´ del Dr. Roderic Guigo´ Serra al Departament de Ciencies` Experimentals i de la Salut de la Universitat Pompeu Fabra Roderic Guigo Serra Sergi Castellano Hereza Barcelona, May 2004 To my parents To my brother and sister Dipòsit legal: B.49649-2004 ISBN: 84-689-0206-3 Preface The election of selenoprotein genes as the subject of a PhD within the area of gene prediction is, and no effort is made here to deny that, a rare case. Not only because it focus on particular type of genes scarcely present in the genome, but because these genes subvert some biological rules we all trust and carefully implement in our gene finding schemas. However, this dissertation and, in general this line of research on selenoproteins, is not the result of a random turn or a crazy night promise1. On the contrary, it is originally rooted on a very specific question posed by previous experimental results on selenoproteins in the fly model system and, not negligible, on the belief that the study of exceptions, if so, can provide great insight into biology. These works in Drosophila were initiated by Montserrat Corominas and Florenci Serras at the Uni- versitat de Barcelona.
    [Show full text]