Mutually exclusive locales for N-linked glycans and disorder in glycoproteins

AbstractSeveral post-translational modifications of proteins lie within regions of disorder, stretches of amino acid residues that exhibit a dynamic tertiary structure and resist crystallization. Such localization has been proposed to expand the binding versatility of the disordered regions, and hence, the repertoire of interacting partners for the proteins. However, investigating a dataset of 500 human N-linked glycoproteins, we observed that the sites of N-linked glycosylations, or N-glycosites, lay predominantly within the regions of predicted order rather than their unstructured counterparts. This mutual exclusivity between disordered stretches and N-glycosites could not be reconciled merely through asymmetry in distribution of asparagines, serines or threonines residues, which comprise the minimum-required signature for conjugation by N-linked glycans, but rather by a contextual enrichment of these residues next to each other within the ordered portions. In fact, N-glycosite neighborhoods and disordered stretches showed distinct sets of enriched residues suggesting their individualized roles in protein phenotype. N-glycosite neighborhood residues also showed higher phylogenetic conservation than disordered stretches within amniote orthologs of glycoproteins. However, a universal search for residue-combinations that are putatively domain-constitutive ranked the disordered regions higher than the N-glycosite neighborhoods. We propose that amino acid residue-combinations bias the permissivity for N-glycoconjugation within ordered regions, so as to balance the tradeoff between the evolution of protein stability, and function, contributed by the N-linked glycans and disordered regions respectively.

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Regulation of N-linked core glycosylation: use of a site-directed mutagenesis approach to identify Asn-Xaa-Ser/Thr sequons that are poor oligosaccharide acceptors

Biochemical Journal ◽

10.1042/bj3230415 ◽

1997 ◽

Vol 323 (2) ◽

pp. 415-419 ◽

Cited By ~ 94

Author(s):

Lakshmi KASTURI ◽

Hegang CHEN ◽

Susan H. SHAKIN-ESHLEMAN

Keyword(s):

Amino Acid ◽

Amino Acid Residue ◽

Site Directed Mutagenesis ◽

Directed Mutagenesis ◽

Amino Acid Residues ◽

Free Translation ◽

A Cell ◽

And Function ◽

A Site ◽

The Impact

N-linked glycosylation can profoundly affect protein expression and function. N-linked glycosylation usually occurs at the sequon Asn-Xaa-Ser/Thr, where Xaa is any amino acid residue except Pro. However, many Asn-Xaa-Ser/Thr sequons are glycosylated inefficiently or not at all for reasons that are poorly understood. We have used a site-directed mutagenesis approach to examine how the Xaa and hydroxy (Ser/Thr) amino acid residues in sequons influence core-glycosylation efficiency. We recently demonstrated that certain Xaa amino acids inhibit core glycosylation of the sequon, Asn37-Xaa-Ser, in rabies virus glycoprotein (RGP). Here we examine the impact of different Xaa residues on core-glycosylation efficiency when the Ser residue in this sequon is replaced with Thr. The core-glycosylation efficiencies of RGP variants with different Asn37-Xaa-Ser/Thr sequons were compared by using a cell-free translation/glycosylation system. Using this approach we confirm that four Asn-Xaa-Ser sequons are poor oligosaccharide acceptors: Asn-Trp-Ser, Asn-Asp-Ser, Asn-Glu-Ser and Asn-Leu-Ser. In contrast, Asn-Xaa-Thr sequons are efficiently glycosylated, even when Xaa = Trp, Asp, Glu or Leu. A comparison of the glycosylation status of Asn-Xaa-Ser and Asn-Xaa-Thr sequons in other glycoproteins confirms that sequons with Xaa = Trp, Asp, Glu or Leu are rarely glycosylated when Ser is the hydroxy amino acid residue, and that these sequons are unlikely to serve as glycosylation sites when introduced into proteins by site-directed mutagenesis.

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Identification of Amino Acid Residues within the N-Terminal Domain of EspA That Play a Role in EspA Filament Biogenesis and Function

Journal of Bacteriology ◽

10.1128/jb.01753-07 ◽

2008 ◽

Vol 190 (6) ◽

pp. 2221-2226 ◽

Cited By ~ 5

Author(s):

Mona P. Singh ◽

Robert K. Shaw ◽

Stuart Knutton ◽

Mark J. Pallen ◽

Valerie F. Crepin ◽

...

Keyword(s):

Escherichia Coli ◽

Amino Acid ◽

Protein Stability ◽

Type Iii Secretion ◽

Type Iii Secretion System ◽

Secretion System ◽

Terminal Region ◽

Translocator Protein ◽

Amino Acid Residues ◽

And Function

ABSTRACT Enteropathogenic Escherichia coli employs a filamentous type III secretion system, made by homopolymerization of the translocator protein EspA. In this study, we have shown that the N-terminal region of EspA has a role in EspA's protein stability, interaction with the CesAB chaperone, and filament biogenesis and function.

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Effect of roasting temperature on lipid and protein oxidation and amino acid residue side chain modification of beef patties

RSC Advances ◽

10.1039/d1ra03151a ◽

2021 ◽

Vol 11 (35) ◽

pp. 21629-21641

Author(s):

Chao Xia ◽

Pingping Wen ◽

Yaming Yuan ◽

Xiaofan Yu ◽

Yijing Chen ◽

...

Keyword(s):

Amino Acid ◽

Amino Acid Residue ◽

Protein Oxidation ◽

Relative Number ◽

Side Chain ◽

Amino Acid Residues ◽

Beef Patties ◽

Lipid And Protein Oxidation ◽

Roasting Temperature

The relative number of peptides modified by the amino acid residues of actin from raw beef patties and those cooked at different roasting temperatures.

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Chitosanase from Streptomyces sp. strain N174: a comparative review of its structure and function

Biochemistry and Cell Biology ◽

10.1139/o97-079 ◽

1997 ◽

Vol 75 (6) ◽

pp. 687-696 ◽

Cited By ~ 20

Author(s):

Tamo Fukamizo ◽

Ryszard Brzezinski

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Substrate Binding ◽

Structure And Function ◽

Site Directed Mutagenesis ◽

Directed Mutagenesis ◽

Amino Acid Residues ◽

Streptomyces Sp ◽

Binding Cleft ◽

And Function

Novel information on the structure and function of chitosanase, which hydrolyzes the beta -1,4-glycosidic linkage of chitosan, has accumulated in recent years. The cloning of the chitosanase gene from Streptomyces sp. strain N174 and the establishment of an efficient expression system using Streptomyces lividans TK24 have contributed to these advances. Amino acid sequence comparisons of the chitosanases that have been sequenced to date revealed a significant homology in the N-terminal module. From energy minimization based on the X-ray crystal structure of Streptomyces sp. strain N174 chitosanase, the substrate binding cleft of this enzyme was estimated to be composed of six monosaccharide binding subsites. The hydrolytic reaction takes place at the center of the binding cleft with an inverting mechanism. Site-directed mutagenesis of the carboxylic amino acid residues that are conserved revealed that Glu-22 and Asp-40 are the catalytic residues. The tryptophan residues in the chitosanase do not participate directly in the substrate binding but stabilize the protein structure by interacting with hydrophobic and carboxylic side chains of the other amino acid residues. Structural and functional similarities were found between chitosanase, barley chitinase, bacteriophage T4 lysozyme, and goose egg white lysozyme, even though these proteins share no sequence similarities. This information can be helpful for the design of new chitinolytic enzymes that can be applied to carbohydrate engineering, biological control of phytopathogens, and other fields including chitinous polysaccharide degradation. Key words: chitosanase, amino acid sequence, overexpression system, reaction mechanism, site-directed mutagenesis.

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Evolutionary divergence and salinity-mediated selection in halophilic archaea

Microbiology and Molecular Biology Reviews ◽

10.1128/mmbr.61.1.90-104.1997 ◽

1997 ◽

Vol 61 (1) ◽

pp. 90-104

Author(s):

P P Dennis ◽

L C Shimmin

Keyword(s):

Amino Acid ◽

Tertiary Structure ◽

Halophilic Archaea ◽

Amino Acid Replacement ◽

Evolutionary Divergence ◽

Ionic Balance ◽

Amino Acid Residues ◽

Nucleotide Substitutions ◽

Nonsynonymous Substitutions ◽

Environmental Salinity

Halophilic (literally salt-loving) archaea are a highly evolved group of organisms that are uniquely able to survive in and exploit hypersaline environments. In this review, we examine the potential interplay between fluctuations in environmental salinity and the primary sequence and tertiary structure of halophilic proteins. The proteins of halophilic archaea are highly adapted and magnificently engineered to function in an intracellular milieu that is in ionic balance with an external environment containing between 2 and 5 M inorganic salt. To understand the nature of halophilic adaptation and to visualize this interplay, the sequences of genes encoding the L11, L1, L10, and L12 proteins of the large ribosome subunit and Mn/Fe superoxide dismutase proteins from three genera of halophilic archaea have been aligned and analyzed for the presence of synonymous and nonsynonymous nucleotide substitutions. Compared to homologous eubacterial genes, these halophilic genes exhibit an inordinately high proportion of nonsynonymous nucleotide substitutions that result in amino acid replacement in the encoded proteins. More than one-third of the replacements involve acidic amino acid residues. We suggest that fluctuations in environmental salinity provide the driving force for fixation of the excessive number of nonsynonymous substitutions. Tinkering with the number, location, and arrangement of acidic and other amino acid residues influences the fitness (i.e., hydrophobicity, surface hydration, and structural stability) of the halophilic protein. Tinkering is also evident at halophilic protein positions monomorphic or polymorphic for serine; more than one-third of these positions use both the TCN and the AGY serine codons, indicating that there have been multiple nonsynonymous substitutions at these positions. Our model suggests that fluctuating environmental salinity prevents optimization of fitness for many halophilic proteins and helps to explain the unusual evolutionary divergence of their encoding genes.

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Identification and Analysis of Novel Amino-Acid Sequence Repeats inBacillus anthracisstr.AmesProteome Using Computational Tools

Comparative and Functional Genomics ◽

10.1155/2007/47161 ◽

2007 ◽

Vol 2007 ◽

pp. 1-23 ◽

Cited By ~ 2

Author(s):

G. R. Hemalatha ◽

D. Satyanarayana Rao ◽

L. Guruprasad

Keyword(s):

Amino Acid ◽

Amino Acid Residue ◽

Protein Sequence ◽

Amino Acid Residues ◽

Sequence Motifs ◽

Computational Tools ◽

Conserved Sequence ◽

Conserved Sequence Motifs ◽

Multiple Copies ◽

Domain 3

We have identified four repeats and ten domains that are novel in proteins encoded by theBacillus anthracisstr.Amesproteome using automated in silico methods. A “repeat” corresponds to a region comprising less than 55-amino-acid residues that occur more than once in the protein sequence and sometimes present in tandem. A “domain” corresponds to a conserved region with greater than 55-amino-acid residues and may be present as single or multiple copies in the protein sequence. These correspond to (1) 57-amino-acid-residue PxV domain, (2) 122-amino-acid-residue FxF domain, (3) 111-amino-acid-residue YEFF domain, (4) 109-amino-acid-residue IMxxH domain, (5) 103-amino-acid-residue VxxT domain, (6) 84-amino-acid-residue ExW domain, (7) 104-amino-acid-residue NTGFIG domain, (8) 36-amino-acid-residue NxGK repeat, (9) 95-amino-acid-residue VYV domain, (10) 75-amino-acid-residue KEWE domain, (11) 59-amino-acid-residue AFL domain, (12) 53-amino-acid-residue RIDVK repeat, (13) (a) 41-amino-acid-residue AGQF repeat and (b) 42-amino-acid-residue GSAL repeat. A repeat or domain type is characterized by specific conserved sequence motifs. We discuss the presence of these repeats and domains in proteins from other genomes and their probable secondary structure.

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Synthesis and Evaluation of Cyclic Acetals of Serine Hydroxylamine for Amide-Forming KAHA Ligations

Synthesis ◽

10.1055/s-0037-1611635 ◽

2019 ◽

Vol 51 (05) ◽

pp. 1273-1283 ◽

Cited By ~ 1

Author(s):

Simon Baldauf ◽

Jeffrey Bode

Keyword(s):

Amino Acid ◽

Amino Acid Residue ◽

Amino Acid Residues ◽

High Demand ◽

Cyclic Acetals ◽

Ligation Product ◽

Canonical Amino Acid ◽

The Stability ◽

Peptide Segments ◽

Derivatives Of

The α-ketoacid–hydroxylamine (KAHA) ligation allows the coupling of unprotected peptide segments. The most widely used variant employs a 5-membered cyclic hydroxylamine that forms a homoserine ester as the primary ligation product. While very effective, monomers that give canonical amino acid residues are in high demand. In order to preserve the stability and reactivity of cyclic hydroxylamines, but form a canonical amino acid residue upon ligation, we sought to prepare cyclic derivatives of serine hydroxylamine. An evaluation of several cyclization strategies led to cyclobutanone ketals as the leading structures. The preparation, stability, and amide-forming ligation of these serine-derived ketals are described.

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Synthesis of arborane triterpenols by a bacterial oxidosqualene cyclase

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1617231114 ◽

2016 ◽

Vol 114 (2) ◽

pp. 245-250 ◽

Cited By ~ 17

Author(s):

Amy B. Banta ◽

Jeremy H. Wei ◽

Clare C. C. Gill ◽

José-Luis Giner ◽

Paula V. Welander

Keyword(s):

Amino Acid ◽

Broad Class ◽

Heterotrophic Bacterium ◽

Expression System ◽

Amino Acid Residues ◽

Biologically Relevant ◽

Oxidosqualene Cyclase ◽

Membrane Structure And Function ◽

Terrestrial Input ◽

And Function

Cyclic triterpenoids are a broad class of polycyclic lipids produced by bacteria and eukaryotes. They are biologically relevant for their roles in cellular physiology, including membrane structure and function, and biochemically relevant for their exquisite enzymatic cyclization mechanism. Cyclic triterpenoids are also geobiologically significant as they are readily preserved in sediments and are used as biomarkers for ancient life throughout Earth's history. Isoarborinol is one such triterpenoid whose only known biological sources are certain angiosperms and whose diagenetic derivatives (arboranes) are often used as indicators of terrestrial input into aquatic environments. However, the occurrence of arborane biomarkers in Permian and Triassic sediments, which predates the accepted origin of angiosperms, suggests that microbial sources of these lipids may also exist. In this study, we identify two isoarborinol-like lipids, eudoraenol and adriaticol, produced by the aerobic marine heterotrophic bacterium Eudoraea adriatica. Phylogenetic analysis demonstrates that the E. adriatica eudoraenol synthase is an oxidosqualene cyclase homologous to bacterial lanosterol synthases and distinct from plant triterpenoid synthases. Using an Escherichia coli heterologous sterol expression system, we demonstrate that substitution of four amino acid residues in a bacterial lanosterol synthase enabled synthesis of pentacyclic arborinols in addition to tetracyclic sterols. This variant provides valuable mechanistic insight into triterpenoid synthesis and reveals diagnostic amino acid residues to differentiate between sterol and arborinol synthases in genomic and metagenomic datasets. Our data suggest that there may be additional bacterial arborinol producers in marine and freshwater environments that could expand our understanding of these geologically informative lipids.

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Identification and characterization of two types of amino acid-regulated acetyltransferases in actinobacteria

Bioscience Reports ◽

10.1042/bsr20170157 ◽

2017 ◽

Vol 37 (4) ◽

Cited By ~ 5

Author(s):

Yu-Xing Lu ◽

Xin-Xin Liu ◽

Wei-Bing Liu ◽

Bang-Ce Ye

Keyword(s):

Amino Acid ◽

Feedback Regulation ◽

Protein Acetylation ◽

Post Translational Modifications ◽

Act Domain ◽

Amino Acid Binding ◽

Amino Acid Sensing ◽

And Function ◽

Future Work ◽

The Relationship

Abstract One hundred and fifty GCN5-like acetyltransferases with amino acid-binding (ACT)-GCN5-related N-acetyltransferase (GNAT) domain organization have been identified in actinobacteria. The ACT domain is fused to the GNAT domain, conferring amino acid-induced allosteric regulation to these protein acetyltransferases (Pat) (amino acid sensing acetyltransferase, (AAPatA)). Members of the AAPatA family share similar secondary structure and are divided into two groups based on the allosteric ligands of the ACT domain: the asparagine (Asn)-activated PatA and the cysteine (Cys)-activated PatA. The former are mainly found in Streptomyces; the latter are distributed in other actinobacteria. We investigated the effect of Asn and Cys on the acetylation activity of Sven_0867 (SvePatA, from Streptomyces venezuelae DSM 40230) and Amir_5672 (AmiPatA, from Actinosynnema mirum strain DSM 43827), respectively, as well as the relationship between the structure and function of these enzymes. These findings indicate that the activity of PatA and acetylation level of proteins may be closely correlated with intracellular concentrations of Asn and Cys in actinobacteria. Amino acid-sensing signal transduction in acetyltransferases may be a mechanism that regulates protein acetylation in response to nutrient availability. Future work examining the relationship between protein acetylation and amino acid metabolism will broaden our understanding of post-translational modifications (PTMs) in feedback regulation.

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