Aharoni A


Full name : Aharoni Amir

First name : Amir

Mail : Department of Life Sciences National Institute for Biotechnology in the Negev, Be'er Sheva 84105

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Country : Israel

Email : aaharoni@bgu.ac.il

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References (12)

Title : The Plant Cuticle: An Ancient Guardian Barrier Set Against Long-Standing Rivals - Arya_2021_Front.Plant.Sci_12_663165
Author(s) : Arya GC , Sarkar S , Manasherova E , Aharoni A , Cohen H
Ref : Front Plant Sci , 12 :663165 , 2021
Abstract : The aerial surfaces of plants are covered by a protective barrier formed by the cutin polyester and waxes, collectively referred to as the cuticle. Plant cuticles prevent the loss of water, regulate transpiration, and facilitate the transport of gases and solutes. As the cuticle covers the outermost epidermal cell layer, it also acts as the first line of defense against environmental cues and biotic stresses triggered by a large array of pathogens and pests, such as fungi, bacteria, and insects. Numerous studies highlight the cuticle interface as the site of complex molecular interactions between plants and pathogens. Here, we outline the multidimensional roles of cuticle-derived components, namely, epicuticular waxes and cutin monomers, during plant interactions with pathogenic fungi. We describe how certain wax components affect various pre-penetration and infection processes of fungi with different lifestyles, and then shift our focus to the roles played by the cutin monomers that are released from the cuticle owing to the activity of fungal cutinases during the early stages of infection. We discuss how cutin monomers can activate fungal cutinases and initiate the formation of infection organs, the significant impacts of cuticle defects on the nature of plant-fungal interactions, along with the possible mechanisms raised thus far in the debate on how host plants perceive cutin monomers and/or cuticle defects to elicit defense responses.
ESTHER : Arya_2021_Front.Plant.Sci_12_663165
PubMedSearch : Arya_2021_Front.Plant.Sci_12_663165
PubMedID: 34249035

Title : Automated Structure- and Sequence-Based Design of Proteins for High Bacterial Expression and Stability - Goldenzweig_2016_Mol.Cell_63_337
Author(s) : Goldenzweig A , Goldsmith M , Hill SE , Gertman O , Laurino P , Ashani Y , Dym O , Unger T , Albeck S , Prilusky J , Lieberman RL , Aharoni A , Silman I , Sussman JL , Tawfik DS , Fleishman SJ
Ref : Mol Cell , 63 :337 , 2016
Abstract : Upon heterologous overexpression, many proteins misfold or aggregate, thus resulting in low functional yields. Human acetylcholinesterase (hAChE), an enzyme mediating synaptic transmission, is a typical case of a human protein that necessitates mammalian systems to obtain functional expression. We developed a computational strategy and designed an AChE variant bearing 51 mutations that improved core packing, surface polarity, and backbone rigidity. This variant expressed at approximately 2,000-fold higher levels in E. coli compared to wild-type hAChE and exhibited 20 degrees C higher thermostability with no change in enzymatic properties or in the active-site configuration as determined by crystallography. To demonstrate broad utility, we similarly designed four other human and bacterial proteins. Testing at most three designs per protein, we obtained enhanced stability and/or higher yields of soluble and active protein in E. coli. Our algorithm requires only a 3D structure and several dozen sequences of naturally occurring homologs, and is available at http://pross.weizmann.ac.il.
ESTHER : Goldenzweig_2016_Mol.Cell_63_337
PubMedSearch : Goldenzweig_2016_Mol.Cell_63_337
PubMedID: 27425410
Gene_locus related to this paper: human-ACHE

Title : Overexpression of AtSHN1\/WIN1 Provokes Unique Defense Responses - Sela_2013_PLoS.One_8_e70146
Author(s) : Sela D , Buxdorf K , Shi JX , Feldmesser E , Schreiber L , Aharoni A , Levy M
Ref : PLoS ONE , 8 :e70146 , 2013
Abstract : The plant cell cuticle serves as the first barrier protecting plants from mechanical injury and invading pathogens. The cuticle can be breached by cutinase-producing pathogens and the degradation products may activate pathogenesis signals in the invading pathogens. Cuticle degradation products may also trigger the plant's defense responses. Botrytis cinerea is an important plant pathogen, capable of attacking and causing disease in a wide range of plant species. Arabidopsis thaliana shn1-1D is a gain-of-function mutant, which has a modified cuticular lipid composition. We used this mutant to examine the effect of altering the whole-cuticle metabolic pathway on plant responses to B. cinerea attack. Following infection with B. cinerea, the shn1-1D mutant discolored more quickly, accumulated more H2O2, and showed accelerated cell death relative to wild-type (WT) plants. Whole transcriptome analysis of B. cinerea-inoculated shn1-1D vs. WT plants revealed marked upregulation of genes associated with senescence, oxidative stress and defense responses on the one hand, and genes involved in the magnitude of defense-response control on the other. We propose that altered cutin monomer content and composition of shn1-1D plants triggers excessive reactive oxygen species accumulation and release which leads to a strong, unique and uncontrollable defense response, resulting in plant sensitivity and death.
ESTHER : Sela_2013_PLoS.One_8_e70146
PubMedSearch : Sela_2013_PLoS.One_8_e70146
PubMedID: 23922943

Title : Employing directed evolution for the functional analysis of multi-specific proteins - Levin_2013_Bioorg.Med.Chem_21_3511
Author(s) : Levin M , Amar D , Aharoni A
Ref : Bioorganic & Medicinal Chemistry , 21 :3511 , 2013
Abstract : Multi-specific proteins located at the heart of complex protein-protein interaction (PPI) networks play essential roles in the survival and fitness of the cell. In addition, multi-specific or promiscuous enzymes exhibit activity toward a wide range of substrates so as to increase cell evolvability and robustness. However, despite their high importance, investigating the in vivo function of these proteins is difficult, due to their complex nature. Typically, deletion of these proteins leads to the abolishment of large PPI networks, highlighting the difficulty in examining the contributions of specific interactions/activities to complex biological processes and cell phenotypes. Protein engineering approaches, including directed evolution and computational protein design, allow for the generation of multi-specific proteins in which certain activities remain intact while others are abolished. The generation and examination of these mutants both in vitro and in vivo can provide high-resolution analysis of biological processes and cell phenotypes and provide new insight into the evolution and molecular function of this important protein family.
ESTHER : Levin_2013_Bioorg.Med.Chem_21_3511
PubMedSearch : Levin_2013_Bioorg.Med.Chem_21_3511
PubMedID: 23683833

Title : Novel hybrid esterase-haloacid dehalogenase enzyme -
Author(s) : Beloqui A , Polaina J , Vieites JM , Reyes-Duarte D , Torres R , Golyshina OV , Chernikova TN , Waliczek A , Aharoni A , Yakimov MM , Timmis KN , Golyshin PN , Ferrer M
Ref : Chembiochem , 11 :1975 , 2010
PubMedID: 20715265

Title : High-throughput screening of enzyme libraries: thiolactonases evolved by fluorescence-activated sorting of single cells in emulsion compartments - Aharoni_2005_Chem.Biol_12_1281
Author(s) : Aharoni A , Amitai G , Bernath K , Magdassi S , Tawfik DS
Ref : Chemical Biology , 12 :1281 , 2005
Abstract : Single bacterial cells, each expressing a different library variant, were compartmentalized in aqueous droplets of water-in-oil (w/o) emulsions, thus maintaining a linkage between a plasmid-borne gene, the encoded enzyme variant, and the fluorescent product this enzyme may generate. Conversion into a double, water-in-oil-in-water (w/o/w) emulsion enabled the sorting of these compartments by FACS, as well as the isolation of living bacteria cells and their enzyme-coding genes. We demonstrate the directed evolution of new enzyme variants by screening >10(7) serum paraoxonase (PON1) mutants, to yield 100-fold improvements in thiolactonase activity. In vitro compartmentalization (IVC) of single cells, each carrying >10(4) enzyme molecules, in a volume of <10 femtoliter (fl), enabled detection and selection despite the fast, spontaneous hydrolysis of the substrate, the very low initial thiolactonase activity of PON1, and the use of difusable fluorescent products.
ESTHER : Aharoni_2005_Chem.Biol_12_1281
PubMedSearch : Aharoni_2005_Chem.Biol_12_1281
PubMedID: 16356845

Title : The 'evolvability' of promiscuous protein functions - Aharoni_2005_Nat.Genet_37_73
Author(s) : Aharoni A , Gaidukov L , Khersonsky O , Mc QGS , Roodveldt C , Tawfik DS
Ref : Nat Genet , 37 :73 , 2005
Abstract : How proteins with new functions (e.g., drug or antibiotic resistance or degradation of man-made chemicals) evolve in a matter of months or years is still unclear. This ability is dependent on the induction of new phenotypic traits by a small number of mutations (plasticity). But mutations often have deleterious effects on functions that are essential for survival. How are these seemingly conflicting demands met at the single-protein level? Results from directed laboratory evolution experiments indicate that the evolution of a new function is driven by mutations that have little effect on the native function but large effects on the promiscuous functions that serve as starting point. Thus, an evolving protein can initially acquire increased fitness for a new function without losing its original function. Gene duplication and the divergence of a completely new protein may then follow.
ESTHER : Aharoni_2005_Nat.Genet_37_73
PubMedSearch : Aharoni_2005_Nat.Genet_37_73
PubMedID: 15568024

Title : High-throughput screens and selections of enzyme-encoding genes - Aharoni_2005_Curr.Opin.Chem.Biol_9_210
Author(s) : Aharoni A , Griffiths AD , Tawfik DS
Ref : Curr Opin Chemical Biology , 9 :210 , 2005
Abstract : The availability of vast gene repertoires from both natural sources (genomic and cDNA libraries) and artificial sources (gene libraries) demands the development and application of novel technologies that enable the screening or selection of large libraries for a variety of enzymatic activities. We describe recent developments in the selection of enzyme-coding genes for directed evolution and functional genomics. We focus on HTS approaches that enable selection from large libraries (>10(6) gene variants) with relatively humble means (i.e. non-robotic systems), and on in vitro compartmentalization in particular.
ESTHER : Aharoni_2005_Curr.Opin.Chem.Biol_9_210
PubMedSearch : Aharoni_2005_Curr.Opin.Chem.Biol_9_210
PubMedID: 15811807

Title : Directed evolution of proteins for heterologous expression and stability - Roodveldt_2005_Curr.Opin.Struct.Biol_15_50
Author(s) : Roodveldt C , Aharoni A , Tawfik DS
Ref : Current Opinion in Structural Biology , 15 :50 , 2005
Abstract : Recent developments have been made in the application of directed evolution to achieve the efficient heterologous expression of proteins in Escherichia coli and yeast by increasing the stability and solubility of the protein in the host environment. One interesting conclusion that emerges is that the evolutionary process often improves the stability and solubility of an intermediate (apoprotein, proprotein or folding intermediate) that otherwise constitutes a bottleneck to functional expression, rather than altering the protein's final state.
ESTHER : Roodveldt_2005_Curr.Opin.Struct.Biol_15_50
PubMedSearch : Roodveldt_2005_Curr.Opin.Struct.Biol_15_50
PubMedID: 15718133

Title : Structure and evolution of the serum paraoxonase family of detoxifying and anti-atherosclerotic enzymes - Harel_2004_Nat.Struct.Mol.Biol_11_412
Author(s) : Harel M , Aharoni A , Gaidukov L , Brumshtein B , Khersonsky O , Meged R , Dvir H , Ravelli RB , McCarthy A , Toker L , Silman I , Sussman JL , Tawfik DS
Ref : Nat Struct Mol Biol , 11 :412 , 2004
Abstract : Members of the serum paraoxonase (PON) family have been identified in mammals and other vertebrates, and in invertebrates. PONs exhibit a wide range of physiologically important hydrolytic activities, including drug metabolism and detoxification of nerve agents. PON1 and PON3 reside on high-density lipoprotein (HDL, 'good cholesterol') and are involved in the prevention of atherosclerosis. We describe the first crystal structure of a PON family member, a variant of PON1 obtained by directed evolution, at a resolution of 2.2 A. PON1 is a six-bladed beta-propeller with a unique active site lid that is also involved in HDL binding. The three-dimensional structure and directed evolution studies permit a detailed description of PON1's active site and catalytic mechanism, which are reminiscent of secreted phospholipase A2, and of the routes by which PON family members diverged toward different substrate and reaction selectivities.
ESTHER : Harel_2004_Nat.Struct.Mol.Biol_11_412
PubMedSearch : Harel_2004_Nat.Struct.Mol.Biol_11_412
PubMedID: 15098021

Title : Directed evolution of mammalian paraoxonases PON1 and PON3 for bacterial expression and catalytic specialization - Aharoni_2004_Proc.Natl.Acad.Sci.U.S.A_101_482
Author(s) : Aharoni A , Gaidukov L , Yagur S , Toker L , Silman I , Tawfik DS
Ref : Proc Natl Acad Sci U S A , 101 :482 , 2004
Abstract : Serum paraoxonases (PONs) are a group of enzymes that play a key role in organophosphate (OP) detoxification and in prevention of atherosclerosis. However, their structure and mechanism of action are poorly understood. PONs seem like jacks-of-all-trades, acting on a very wide range of substrates, most of which are of no physiological relevance. Family shuffling and screening lead to the first PON variants that express in a soluble and active form in Escherichia coli. We describe variants with kinetic parameters similar to those reported for PONs purified from sera and others that show dramatically increased activities. In particular, we have evolved PON1 variants with OP-hydrolyzing activities 40-fold higher than wild type and a specificity switch of >2,000-fold, producing PONs specialized for OP rather than ester hydrolysis. Analysis of the newly evolved variants provides insights into the evolutionary relationships between different family members.
ESTHER : Aharoni_2004_Proc.Natl.Acad.Sci.U.S.A_101_482
PubMedSearch : Aharoni_2004_Proc.Natl.Acad.Sci.U.S.A_101_482
PubMedID: 14695884

Title : Directed evolution of recombinant serum paraoxonase (PON) variants - Aharoni_2004_Discov.Med_4_120
Author(s) : Aharoni A , Tawfik DS
Ref : Discov Med , 4 :120 , 2004
Abstract : Extract: Owing to their detoxifying functions, and roles in drug metabolism as well as the prevention of atherosclerosis, mammalian or serum paraoxonases (PONs) are an intriguing subject of research and a prime therapeutic and engineering target. Initially identified in mammals, PON and PON-related genes have now been found in fowls, zebra fish, and even in invertebrates such as C. elegans. The more closely-related PON genes are divided into three classes or sub-families: PON1, PON2 and PON3, that share 60-70% sequence identity. PONs are calcium-dependent hydrolases that catalyze the hydrolysis of a broad range of esters and lactones. PON1, which is by far the most investigated member of this family, also catalyzes, albeit at much lower rates, the hydrolysis and thereby inactivation of various organophosphates (OPs), including the nerve agents sarin and soman. PON1 is also involved in drug metabolism and is used for drug inactivation. In recent years, it has become apparent that PONs also play an important role in the prevention of atherosclerosis. The levels of PON1 in the blood and its catalytic proficiency appear to have a major impact both on the individual's susceptibility to pollutants and insecticides, and to atherosclerosis. Furthermore, mice lacking the PON1 gene are highly susceptible to atherosclerosis and to OP poisoning. PON1 and PON3 reside in the high-density lipoprotein cholesterol-carrying particles known as HDL ("good cholesterol"). HDL has two key roles: mediation of cholesterol efflux, e.g., from macrophage foam cells in atherosclerotic lesions, and limitation of lipid oxidation in LDL. PONs have been implicated in both activities.
ESTHER : Aharoni_2004_Discov.Med_4_120
PubMedSearch : Aharoni_2004_Discov.Med_4_120
PubMedID: 20705007