The Coupling of SecA Expression to Secretion Efficiency by SecM-Mediated Translation Arrest

2014 ◽  
pp. 241-256
Author(s):  
Harris D. Bernstein
Keyword(s):  
Translation Arrest ◽  
Secretion Efficiency

Influence of nascent polypeptide positive charges on translation dynamics

2020 ◽  
Vol 477 (15) ◽  
pp. 2921-2934
Author(s):  
Rodrigo D. Requião ◽  
Géssica C. Barros ◽  
Tatiana Domitrovic ◽  
Fernando L. Palhano
Keyword(s):  
Mrna Decay ◽  
Translation Termination ◽  
Published Data ◽  
Translation Efficiency ◽  
Amino Acid Residues ◽  
High Concentration ◽  
Strong Argument ◽  
Translation Arrest ◽  
Exit Tunnel ◽  
Positively Charged

Protein segments with a high concentration of positively charged amino acid residues are often used in reporter constructs designed to activate ribosomal mRNA/protein decay pathways, such as those involving nonstop mRNA decay (NSD), no-go mRNA decay (NGD) and the ribosome quality control (RQC) complex. It has been proposed that the electrostatic interaction of the positively charged nascent peptide with the negatively charged ribosomal exit tunnel leads to translation arrest. When stalled long enough, the translation process is terminated with the degradation of the transcript and an incomplete protein. Although early experiments made a strong argument for this mechanism, other features associated with positively charged reporters, such as codon bias and mRNA and protein structure, have emerged as potent inducers of ribosome stalling. We carefully reviewed the published data on the protein and mRNA expression of artificial constructs with diverse compositions as assessed in different organisms. We concluded that, although polybasic sequences generally lead to lower translation efficiency, it appears that an aggravating factor, such as a nonoptimal codon composition, is necessary to cause translation termination events.

scholarly journals Enhancement of the Efficiency of Secretion of Heterologous Lipase in Escherichia coli by Directed Evolution of the ABC Transporter System

2005 ◽  
Vol 71 (7) ◽  
pp. 3468-3474 ◽  
Author(s):  
Gyeong Tae Eom ◽  
Jae Kwang Song ◽  
Jung Hoon Ahn ◽  
Yeon Soo Seo ◽  
Joon Shick Rhee
Keyword(s):  
Escherichia Coli ◽  
Directed Evolution ◽  
Abc Transporter ◽  
Microtiter Plate ◽  
Single Amino Acid ◽  
Wild Type ◽  
E Coli ◽  
Single Amino Acid Change ◽  
Secretion Efficiency

ABSTRACT The ABC transporter (TliDEF) from Pseudomonas fluorescens SIK W1, which mediated the secretion of a thermostable lipase (TliA) into the extracellular space in Escherichia coli, was engineered using directed evolution (error-prone PCR) to improve its secretion efficiency. TliD mutants with increased secretion efficiency were identified by coexpressing the mutated tliD library with the wild-type tliA lipase in E. coli and by screening the library with a tributyrin-emulsified indicator plate assay and a microtiter plate-based assay. Four selected mutants from one round of error-prone PCR mutagenesis, T6, T8, T24, and T35, showed 3.2-, 2.6-, 2.9-, and 3.0-fold increases in the level of secretion of TliA lipase, respectively, but had almost the same level of expression of TliD in the membrane as the strain with the wild-type TliDEF transporter. These results indicated that the improved secretion of TliA lipase was mediated by the transporter mutations. Each mutant had a single amino acid change in the predicted cytoplasmic regions in the membrane domain of TliD, implying that the corresponding region of TliD was important for the improved and successful secretion of the target protein. We therefore concluded that the efficiency of secretion of a heterologous protein in E. coli can be enhanced by in vitro engineering of the ABC transporter.

scholarly journals Nascent SecM Chain Outside the Ribosome Reinforces Translation Arrest

PLoS ONE ◽  
2015 ◽  
Vol 10 (3) ◽  
pp. e0122017 ◽  
Author(s):  
Zhuohao Yang ◽  
Ryo Iizuka ◽  
Takashi Funatsu
Keyword(s):  
Translation Arrest

scholarly journals Dynamics of the context-specific translation arrest by chloramphenicol and linezolid

2019 ◽  
Vol 16 (3) ◽  
pp. 310-317 ◽  
Author(s):  
Junhong Choi ◽  
James Marks ◽  
Jingji Zhang ◽  
Dong-Hua Chen ◽  
Jinfan Wang ◽  
...  
Keyword(s):  
Translation Arrest ◽  
Context Specific

scholarly journals Genome-wide effects of the antimicrobial peptide apidaecin on translation termination in bacteria

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Kyle Mangano ◽  
Tanja Florin ◽  
Xinhao Shao ◽  
Dorota Klepacki ◽  
Irina Chelysheva ◽  
...  
Keyword(s):  
Antimicrobial Peptide ◽  
Stop Codon ◽  
Translation Termination ◽  
Bacterial Cells ◽  
Release Factor ◽  
Translation Arrest ◽  
Genome Wide Analysis ◽  
Genome Wide ◽  
Exit Tunnel ◽  
Unique Mechanism

Biochemical studies suggested that the antimicrobial peptide apidaecin (Api) inhibits protein synthesis by binding in the nascent peptide exit tunnel and trapping the release factor associated with a terminating ribosome. The mode of Api action in bacterial cells had remained unknown. Here genome-wide analysis reveals that in bacteria, Api arrests translating ribosomes at stop codons and causes pronounced queuing of the trailing ribosomes. By sequestering the available release factors, Api promotes pervasive stop codon bypass, leading to the expression of proteins with C-terminal extensions. Api-mediated translation arrest leads to the futile activation of the ribosome rescue systems. Understanding the unique mechanism of Api action in living cells may facilitate the development of new medicines and research tools for genome exploration.

scholarly journals S3440P Substitution in C-Terminal Region of Human Reelin Dramatically Impairs Secretion of Reelin from HEK 293T Cells

2019 ◽  
Author(s):  
Elika Esmaeilzadeh-Gharehdaghi ◽  
Ehsan Razmara ◽  
Amirreza Bitaraf ◽  
Mahdi Mahmoudi ◽  
Masoud Garshasbi
Keyword(s):  
Tandem Repeats ◽  
Point Mutations ◽  
Random Mutagenesis ◽  
In Vitro Study ◽  
Terminal Region ◽  
Main Role ◽  
Mutant Type ◽  
Secretion Efficiency ◽  
Retzius Cells

Background: Reelin is a large extracellular glycoprotein secreted by Cajal–Retzius cells and has a main role during brain development, especially in neuronal migration. Reelin is comprised of N-terminal F-spondin like domain, eight tandem repeats, and a highly conserved basic C-terminal region (CTR). The CTR main role in the secretion of Reelin has been investigated by advertently inducing deletion in whole or a part of this region; however, the role of CTR point mutations on the secretion of Reelin is shrouded in mystery. Materials and Methods: In this study, we performed experimental analyses on a subregion of Human Reelin containing 5th and 6th repeats (R5-R6), a part of 8 th repeat and the CTR which were amplified from cDNA of K562 and HEPG2 cells and cloned into a mammalian expressional plasmid (pVP22/myc-His). Bioinformatics investigation was performed on the CTR at both level of nucleotide and amino acid as well as mutant type. Random mutagenesis by error-prone PCR method was utilized to induce mutation in the CTR. The secretion efficiency of recombinant wild-type and mutant Reelin constructs compared in cell lysate and supernatant isolated from the transiently transfected HEK 293T cells using 6XHistag ELISA method. Results: In-vitro study demonstrated that the CTR alteration(S3440P) leads to impairment of Reelin secretion even after overexpression. Conclusions: Our results indicate that S3440P substitution in highly conserved structure of the CTR has an important effect on Reelin secretion.

In-vivo magnetic resonance imaging studies of mammogenesis in non-pregnant goats treated with exogenous steroids

1991 ◽  
Vol 58 (2) ◽  
pp. 151-157 ◽  
Author(s):  
Paul A. Fowler ◽  
Christopher H. Knight ◽  
Margaret A. Foster
Keyword(s):  
Magnetic Resonance Imaging ◽  
Magnetic Resonance ◽  
Maximum Size ◽  
Mammary Development ◽  
Mammary Tissue ◽  
Secretory Cells ◽  
Resonance Imaging ◽  
Secretion Efficiency ◽  
Cessation Of Treatment

SummaryMammogenesis and lactation were induced in five multiparous, non-pregnant goats by treatment with oestrogen and progesterone for 11 d, followed by dexamethasone for 3 d. Reserpine was administered during the last 5 d. All five goats lactated, although milk yield was less than had been achieved in previous natural lactations. Mammary development was assessed in vivo, using magnetic resonance imaging. Although parenchyma volume increased by more than 6-fold overall, only 25% of this increase occurred during steroid treatment. Most development took place after the cessation of treatment, when milking commenced. Maximum size was not achieved until week 8 of the induced lactation, and was only 70% of normal parenchyma volume. After 18 weeks lactation the activities of three key milk synthetic enzymes were very similar to values previously found in natural lactations, and secretion efficiency (milk production per unit volume of parenchyma) was also similar to that of natural lactations. We conclude that the lower than normal milk yields were associated with incomplete proliferation of mammary tissue, rather than inadequate differentiation of individual secretory cells.

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