scholarly journals Intra-Protein Coevolution Is Increasingly Functional with Greater Proximity to Fertilization

2020 ◽  
Vol 160 (6) ◽  
pp. 295-308
Author(s):  
Marcel Kwiatkowski ◽  
Abdul R. Asif ◽  
Julia Schumacher ◽  
Bertram Brenig ◽  
Hans Zischler ◽  
...  
Keyword(s):  
Amino Acid ◽  
Potential Effect ◽  
Acid Sites ◽  
Cellular Level ◽  
Node Degree ◽  
Entire Body ◽  
Reproductive Proteins ◽  
Sperm Proteins ◽  
Postcopulatory Selection ◽  
And Function

Intramolecular coevolution of amino acid sites has repeatedly been studied to improve predictions on protein structure and function. Thereby, the focus was on bacterial proteins with available crystallographic data. However, intramolecular coevolution has not yet been compared between protein sets along a gradient of functional proximity to fertilization. This is especially true for the potential effect of external selective forces on intraprotein coevolution. In this study, we investigated both aspects in equally sized sets of mammalian proteins representing spermatozoa, testis, entire body, and liver. For coevolutionary analyses, we derived the proportion of covarying sites per protein from amino acid alignments of 10 mammalian orthologues each. In confirmation of the validity of our coevolution proxy, we found positive associations with the nonsynonymous or amino acid substitution rate in all protein sets. However, our coevolution proxy negatively correlated with the number of protein interactants (node degree) in male reproductive protein sets alone. In addition, a negative association of our coevolution proxy with protein hydrophobicity was significant in sperm proteins only. Accordingly, the restrictive effect of protein interactants was most pronounced in male reproductive proteins, and the tendency of sperm proteins to form internal structures decreased the more coevolutionary sites they had. Both aspects illustrate that the share of outward and thus functional coevolution increases with greater proximity to fertilization. We found this conclusion confirmed by additional comparisons within sperm proteins. Thus, sperm proteins with high hydrophobicity had the lowest proportions of covarying sites and, according to gene annotations, localized more frequently to internal cellular structures. They should therefore be less exposed to postcopulatory forms of sexual selection. Their counterparts with low hydrophobicity had larger proportions of covarying sites and more often resided at the cell membrane or were secreted. At the cellular level, they are thus closer to externally induced forces of postcopulatory selection which are known for their potential to increase substitution rates. In addition, we show that the intermediary status of the testicular protein set in correlation analyses is probably due to a special combination of reproductive and somatic involvements.

scholarly journals Mitochondrial Heterogeneity in the Brain at the Cellular Level

1998 ◽  
Vol 18 (3) ◽  
pp. 231-237 ◽  
Author(s):  
Ursula Sonnewald ◽  
Leif Hertz ◽  
Arne Schousboe
Keyword(s):  
Amino Acid ◽  
Current Knowledge ◽  
Cell Types ◽  
Cellular Level ◽  
Mitochondrial Structure ◽  
Cell Type Specific ◽  
Mitochondrial Heterogeneity ◽  
Specific Tissue ◽  
And Function ◽  
The Brain

Classically, compartmentation of glutamate metabolism in the brain is associated with the fact that neurons and glia exhibit distinct differences with regard to metabolism of this amino acid. The recent use of 13C-labeled compounds to study this metabolism in conjunction with the availability of cell type-specific tissue culture modes has led to the notion that such compartmentation may even be present in individual cell types, neurons as well as glia. To better understand and explain this, it is proposed that mitochondrial heterogeneity may exist resulting in tricarboxylic acid cycles with different properties regarding cycling rates and ratio as well as coupling to amino acid biosynthesis, primarily involving glutamate and aspartate. These hypotheses are evaluated in the light of current knowledge about mitochondrial structure and function.

Structure and function at the cellular level

JAMA ◽  
1966 ◽  
Vol 198 (8) ◽  
pp. 815-825 ◽  
Author(s):  
G. E. Palade
Keyword(s):  
Cellular Level ◽  
Structure And Function ◽  
And Function

scholarly journals Pichia pastoris and the Recombinant Human Heterodimeric Amino Acid Transporter 4F2hc-LAT1: From Clone Selection to Pure Protein

2021 ◽  
Vol 4 (3) ◽  
pp. 51
Author(s):  
Satish Kantipudi ◽  
Daniel Harder ◽  
Sara Bonetti ◽  
Dimitrios Fotiadis ◽  
Jean-Marc Jeckelmann
Keyword(s):  
Amino Acid ◽  
Pichia Pastoris ◽  
Protein Complexes ◽  
Amino Acid Transporter ◽  
Amino Acid Transporters ◽  
Expression Host ◽  
Amino Acids And Derivatives ◽  
Heterodimeric Complex ◽  
Acid Transporter ◽  
And Function

Heterodimeric amino acid transporters (HATs) are protein complexes composed of two subunits, a heavy and a light subunit belonging to the solute carrier (SLC) families SLC3 and SLC7. HATs transport amino acids and derivatives thereof across the plasma membrane. The human HAT 4F2hc-LAT1 is composed of the type-II membrane N-glycoprotein 4F2hc (SLC3A2) and the L-type amino acid transporter LAT1 (SLC7A5). 4F2hc-LAT1 is medically relevant, and its dysfunction and overexpression are associated with autism and tumor progression. Here, we provide a general applicable protocol on how to screen for the best membrane transport protein-expressing clone in terms of protein amount and function using Pichia pastoris as expression host. Furthermore, we describe an overexpression and purification procedure for the production of the HAT 4F2hc-LAT1. The isolated heterodimeric complex is pure, correctly assembled, stable, binds the substrate L-leucine, and is thus properly folded. Therefore, this Pichia pastoris-derived recombinant human 4F2hc-LAT1 sample can be used for downstream biochemical and biophysical characterizations.

scholarly journals Fe(2)OG: an integrated HMM profile-based web server to predict and analyze putative non-haem iron(II)- and 2-oxoglutarate-dependent dioxygenase function in protein sequences

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Siddhartha Kundu
Keyword(s):  
Amino Acid ◽  
Water Molecule ◽  
Active Site ◽  
Ferrous Iron ◽  
Web Server ◽  
Protein Sequences ◽  
Diverse Group ◽  
And Function ◽  
Functionally Diverse ◽  
Haem Iron

Abstract Objective Non-haem iron(II)- and 2-oxoglutarate-dependent dioxygenases (i2OGdd), are a taxonomically and functionally diverse group of enzymes. The active site comprises ferrous iron in a hexa-coordinated distorted octahedron with the apoenzyme, 2-oxoglutarate and a displaceable water molecule. Current information on novel i2OGdd members is sparse and relies on computationally-derived annotation schema. The dissimilar amino acid composition and variable active site geometry thereof, results in differing reaction chemistries amongst i2OGdd members. An additional need of researchers is a curated list of sequences with putative i2OGdd function which can be probed further for empirical data. Results This work reports the implementation of $$Fe\left(2\right)OG$$ F e 2 O G , a web server with dual functionality and an extension of previous work on i2OGdd enzymes $$\left(Fe\left(2\right)OG\equiv \{H2OGpred,DB2OG\}\right)$$ F e 2 O G ≡ { H 2 O G p r e d , D B 2 O G } . $$Fe\left(2\right)OG$$ F e 2 O G , in this form is completely revised, updated (URL, scripts, repository) and will strengthen the knowledge base of investigators on i2OGdd biochemistry and function. $$Fe\left(2\right)OG$$ F e 2 O G , utilizes the superior predictive propensity of HMM-profiles of laboratory validated i2OGdd members to predict probable active site geometries in user-defined protein sequences. $$Fe\left(2\right)OG$$ F e 2 O G , also provides researchers with a pre-compiled list of analyzed and searchable i2OGdd-like sequences, many of which may be clinically relevant. $$Fe(2)OG$$ F e ( 2 ) O G , is freely available (http://204.152.217.16/Fe2OG.html) and supersedes all previous versions, i.e., H2OGpred, DB2OG.

scholarly journals Codon harmonization reduces amino acid misincorporation in bacterially expressed P. falciparum proteins and improves their immunogenicity

AMB Express ◽  
2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Neeraja Punde ◽  
Jennifer Kooken ◽  
Dagmar Leary ◽  
Patricia M. Legler ◽  
Evelina Angov
Keyword(s):  
Protein Structure ◽  
Amino Acid ◽  
Codon Usage ◽  
Dna Sequences ◽  
Structural Integrity ◽  
Host Cells ◽  
Loss Of Function ◽  
Species Specific ◽  
And Function ◽  
The Impact

Abstract Codon usage frequency influences protein structure and function. The frequency with which codons are used potentially impacts primary, secondary and tertiary protein structure. Poor expression, loss of function, insolubility, or truncation can result from species-specific differences in codon usage. “Codon harmonization” more closely aligns native codon usage frequencies with those of the expression host particularly within putative inter-domain segments where slower rates of translation may play a role in protein folding. Heterologous expression of Plasmodium falciparum genes in Escherichia coli has been a challenge due to their AT-rich codon bias and the highly repetitive DNA sequences. Here, codon harmonization was applied to the malarial antigen, CelTOS (Cell-traversal protein for ookinetes and sporozoites). CelTOS is a highly conserved P. falciparum protein involved in cellular traversal through mosquito and vertebrate host cells. It reversibly refolds after thermal denaturation making it a desirable malarial vaccine candidate. Protein expressed in E. coli from a codon harmonized sequence of P. falciparum CelTOS (CH-PfCelTOS) was compared with protein expressed from the native codon sequence (N-PfCelTOS) to assess the impact of codon usage on protein expression levels, solubility, yield, stability, structural integrity, recognition with CelTOS-specific mAbs and immunogenicity in mice. While the translated proteins were expected to be identical, the translated products produced from the codon-harmonized sequence differed in helical content and showed a smaller distribution of polypeptides in mass spectra indicating lower heterogeneity of the codon harmonized version and fewer amino acid misincorporations. Substitutions of hydrophobic-to-hydrophobic amino acid were observed more commonly than any other. CH-PfCelTOS induced significantly higher antibody levels compared with N-PfCelTOS; however, no significant differences in either IFN-γ or IL-4 cellular responses were detected between the two antigens.

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

1997 ◽  
Vol 75 (6) ◽  
pp. 687-696 ◽  
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.

scholarly journals Impact of Aldosterone on the Failing Myocardium: Insights from Mitochondria and Adrenergic Receptors Signaling and Function

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1552
Author(s):  
Mariona Guitart-Mampel ◽  
Pedro Urquiza ◽  
Jordana I. Borges ◽  
Anastasios Lymperopoulos ◽  
Maria E. Solesio
Keyword(s):  
Mitochondrial Dysfunction ◽  
G Protein ◽  
Adrenergic Receptor ◽  
Cellular Level ◽  
Cardiac Physiology ◽  
Failing Heart ◽  
Receptor Kinases ◽  
And Function ◽  
The Impact ◽  
G Protein Coupled

The mineralocorticoid aldosterone regulates electrolyte and blood volume homeostasis, but it also adversely modulates the structure and function of the chronically failing heart, through its elevated production in chronic human post-myocardial infarction (MI) heart failure (HF). By activating the mineralocorticoid receptor (MR), a ligand-regulated transcription factor, aldosterone promotes inflammation and fibrosis of the heart, while increasing oxidative stress, ultimately induding mitochondrial dysfunction in the failing myocardium. To reduce morbidity and mortality in advanced stage HF, MR antagonist drugs, such as spironolactone and eplerenone, are used. In addition to the MR, aldosterone can bind and stimulate other receptors, such as the plasma membrane-residing G protein-coupled estrogen receptor (GPER), further complicating it signaling properties in the myocardium. Given the salient role that adrenergic receptor (ARs)—particularly βARs—play in cardiac physiology and pathology, unsurprisingly, that part of the impact of aldosterone on the failing heart is mediated by its effects on the signaling and function of these receptors. Aldosterone can significantly precipitate the well-documented derangement of cardiac AR signaling and impairment of AR function, critically underlying chronic human HF. One of the main consequences of HF in mammalian models at the cellular level is the presence of mitochondrial dysfunction. As such, preventing mitochondrial dysfunction could be a valid pharmacological target in this condition. This review summarizes the current experimental evidence for this aldosterone/AR crosstalk in both the healthy and failing heart, and the impact of mitochondrial dysfunction in HF. Recent findings from signaling studies focusing on MR and AR crosstalk via non-conventional signaling of molecules that normally terminate the signaling of ARs in the heart, i.e., the G protein-coupled receptor-kinases (GRKs), are also highlighted.

scholarly journals Codon-Substitution Models for Heterogeneous Selection Pressure at Amino Acid Sites

Genetics ◽  
2000 ◽  
Vol 155 (1) ◽  
pp. 431-449 ◽  
Author(s):  
Ziheng Yang ◽  
Rasmus Nielsen ◽  
Nick Goldman ◽  
Anne-Mette Krabbe Pedersen
Keyword(s):  
Amino Acid ◽  
Positive Selection ◽  
Selective Pressure ◽  
Acid Sites ◽  
Data Sets ◽  
Protein Coding ◽  
Important Indicator ◽  
Diversifying Selection ◽  
Codon Substitution ◽  
Neutral Mutations

AbstractComparison of relative fixation rates of synonymous (silent) and nonsynonymous (amino acid-altering) mutations provides a means for understanding the mechanisms of molecular sequence evolution. The nonsynonymous/synonymous rate ratio (ω = dN/dS) is an important indicator of selective pressure at the protein level, with ω = 1 meaning neutral mutations, ω < 1 purifying selection, and ω > 1 diversifying positive selection. Amino acid sites in a protein are expected to be under different selective pressures and have different underlying ω ratios. We develop models that account for heterogeneous ω ratios among amino acid sites and apply them to phylogenetic analyses of protein-coding DNA sequences. These models are useful for testing for adaptive molecular evolution and identifying amino acid sites under diversifying selection. Ten data sets of genes from nuclear, mitochondrial, and viral genomes are analyzed to estimate the distributions of ω among sites. In all data sets analyzed, the selective pressure indicated by the ω ratio is found to be highly heterogeneous among sites. Previously unsuspected Darwinian selection is detected in several genes in which the average ω ratio across sites is <1, but in which some sites are clearly under diversifying selection with ω > 1. Genes undergoing positive selection include the β-globin gene from vertebrates, mitochondrial protein-coding genes from hominoids, the hemagglutinin (HA) gene from human influenza virus A, and HIV-1 env, vif, and pol genes. Tests for the presence of positively selected sites and their subsequent identification appear quite robust to the specific distributional form assumed for ω and can be achieved using any of several models we implement. However, we encountered difficulties in estimating the precise distribution of ω among sites from real data sets.

scholarly journals Genetic Relationships in the Toxin-Producing Fungal Endophyte, Alternaria oxytropis Using Polyketide Synthase and Non-Ribosomal Peptide Synthase Genes

2021 ◽  
Vol 7 (7) ◽  
pp. 538
Author(s):  
Rebecca Creamer ◽  
Deana Baucom Hille ◽  
Marwa Neyaz ◽  
Tesneem Nusayr ◽  
Christopher L. Schardl ◽  
...  
Keyword(s):  
Amino Acid ◽  
Polyketide Synthase ◽  
Genetic Relationships ◽  
Protein Sequences ◽  
Fungal Endophyte ◽  
Melanin Synthesis ◽  
Melanin Biosynthesis ◽  
Protein Levels ◽  
Oxytropis Sericea ◽  
And Function

The legume Oxytropis sericea hosts a fungal endophyte, Alternaria oxytropis, which produces secondary metabolites (SM), including the toxin swainsonine. Polyketide synthase (PKS) and non-ribosomal peptide synthase (NRPS) enzymes are associated with biosynthesis of fungal SM. To better understand the origins of the SM, an unannotated genome of A. oxytropis was assessed for protein sequences similar to known PKS and NRPS enzymes of fungi. Contigs exhibiting identity with known genes were analyzed at nucleotide and protein levels using available databases. Software were used to identify PKS and NRPS domains and predict identity and function. Confirmation of sequence for selected gene sequences was accomplished using PCR. Thirteen PKS, 5 NRPS, and 4 PKS-NRPS hybrids were identified and characterized with functions including swainsonine and melanin biosynthesis. Phylogenetic relationships among closest amino acid matches with Alternaria spp. were identified for seven highly conserved PKS and NRPS, including melanin synthesis. Three PKS and NRPS were most closely related to other fungi within the Pleosporaceae family, while five PKS and PKS-NRPS were closely related to fungi in the Pleosporales order. However, seven PKS and PKS-NRPS showed no identity with fungi in the Pleosporales or the class Dothideomycetes, suggesting a different evolutionary origin for those genes.

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