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Pflugers Archiv : European Journal of... Apr 2024The transport of bicarbonate across the enterocyte cell membrane regulates the intracellular as well as the luminal pH and is an essential part of directional fluid... (Review)
Review
The transport of bicarbonate across the enterocyte cell membrane regulates the intracellular as well as the luminal pH and is an essential part of directional fluid movement in the gut. Since the first description of "active" transport of HCO ions against a concentration gradient in the 1970s, the fundamental role of HCO transport for multiple intestinal functions has been recognized. The ion transport proteins have been identified and molecularly characterized, and knockout mouse models have given insight into their individual role in a variety of functions. This review describes the progress made in the last decade regarding novel techniques and new findings in the molecular regulation of intestinal HCO transport in the different segments of the gut. We discuss human diseases with defects in intestinal HCO secretion and potential treatment strategies to increase luminal alkalinity. In the last part of the review, the cellular and organismal mechanisms for acid/base sensing in the intestinal tract are highlighted.
Topics: Animals; Mice; Humans; Bicarbonates; Ion Transport; Enterocytes; Cell Membrane; Bodily Secretions; Hydrogen-Ion Concentration; Cystic Fibrosis Transmembrane Conductance Regulator
PubMed: 38374228
DOI: 10.1007/s00424-024-02914-3 -
Biochimica Et Biophysica Acta.... Oct 2022Secreted proteins contain an N-terminal signal peptide to guide them through the secretion pathway. Once the protein is translocated, the signal peptide is removed by a... (Review)
Review
Secreted proteins contain an N-terminal signal peptide to guide them through the secretion pathway. Once the protein is translocated, the signal peptide is removed by a signal peptidase, such as signal peptidase I. The signal peptide has been extensively studied and reviewed; however, the mature region has not been the focus of review. Here we cover the experimental evidence that highlights the important role of the mature region amino acid residues in both the efficiency and the ability of secreted proteins to be successfully exported via secretion pathways and cleaved by signal peptidase I.
Topics: Amino Acid Sequence; Bacterial Proteins; Biological Transport; Escherichia coli; Protein Sorting Signals
PubMed: 35798072
DOI: 10.1016/j.bbamem.2022.184000 -
Journal of Bacteriology Mar 2021Large clostridial toxins (LCTs) are secreted virulence factors found in several species, including , , , and LCTs are large toxins that lack a secretion signal...
Large clostridial toxins (LCTs) are secreted virulence factors found in several species, including , , , and LCTs are large toxins that lack a secretion signal sequence, and studies by others have shown that the LCTs of , TcdA and TcdB, require a holin-like protein, TcdE, for secretion. The TcdE gene is located on the pathogenicity locus (PaLoc) of , and holin-encoding genes are also present in the LCT-encoded PaLocs from and However, the holin (TpeE) associated with the LCT TpeL has no homology and a different membrane topology than TcdE. In addition, TpeE has a membrane topology identical to that of the TatA protein, which is the core of the twin-arginine translocation (Tat) secretion system. To determine if TpeE was necessary and sufficient to secrete TpeL, the genes from a type C strain of were expressed in a type A strain of , HN13, and secretion was measured using Western blot methods. We found that TpeE was required for TpeL secretion and that secretion was not due to cell lysis. Mutant forms of TpeE lacking an amphipathic helix and a charged C-terminal domain failed to secrete TpeL, and mutations that deleted conserved LCT domains in TpeL indicated that only the full-length protein could be secreted. In summary, we have identified a novel family of holin-like proteins that can function, in some cases, as a system of protein secretion for proteins that need to fold in the cytoplasm. Little is known about the mechanism by which LCTs are secreted. Since LCTs are major virulence factors in clostridial pathogens, we wanted to define the mechanism by which an LCT in , TpeL, is secreted by a protein (TpeE) lacking homology to previously described secretion-associated holins. We discovered that TpeE is a member of a widely dispersed class of holin proteins, and TpeE is necessary for the secretion of TpeL. TpeE bears a high degree of similarity in membrane topology to TatA proteins, which form the pore through which Tat secretion substrates pass through the cytoplasmic membrane. Thus, the TpeE-TpeL secretion system may be a model for understanding not only holin-dependent secretion but also how TatA proteins function in the secretion process.
Topics: Bacterial Proteins; Bacterial Secretion Systems; Bacterial Toxins; Clostridium perfringens; Gene Expression Regulation, Bacterial; Protein Transport
PubMed: 33526612
DOI: 10.1128/JB.00580-20 -
Biotechnology and Bioengineering Feb 2021Despite their therapeutic potential, many protein drugs remain inaccessible to patients since they are difficult to secrete. Each recombinant protein has unique...
Despite their therapeutic potential, many protein drugs remain inaccessible to patients since they are difficult to secrete. Each recombinant protein has unique physicochemical properties and requires different machinery for proper folding, assembly, and posttranslational modifications (PTMs). Here we aimed to identify the machinery supporting recombinant protein secretion by measuring the protein-protein interaction (PPI) networks of four different recombinant proteins (SERPINA1, SERPINC1, SERPING1, and SeAP) with various PTMs and structural motifs using the proximity-dependent biotin identification (BioID) method. We identified PPIs associated with specific features of the secreted proteins using a Bayesian statistical model and found proteins involved in protein folding, disulfide bond formation, and N-glycosylation were positively correlated with the corresponding features of the four model proteins. Among others, oxidative folding enzymes showed the strongest association with disulfide bond formation, supporting their critical roles in proper folding and maintaining the ER stability. Knockdown of disulfide-isomerase PDIA4, a measured interactor with significance for SERPINC1 but not SERPINA1, led to the decreased secretion of SERPINC1, which relies on its extensive disulfide bonds, compared to SERPINA1, which has no disulfide bonds. Proximity-dependent labeling successfully identified the transient interactions supporting synthesis of secreted recombinant proteins and refined our understanding of key molecular mechanisms of the secretory pathway during recombinant protein production.
Topics: Glycosylation; HEK293 Cells; Humans; Protein Folding; Protein Interaction Maps; Protein Processing, Post-Translational; Protein Transport; Recombinant Proteins
PubMed: 33169829
DOI: 10.1002/bit.27621 -
Trends in Microbiology Nov 2021Efficient penetration of the mucus layer is needed for respiratory viruses to avoid mucociliary clearance prior to infection. Many respiratory viruses bind to glycans on... (Review)
Review
Efficient penetration of the mucus layer is needed for respiratory viruses to avoid mucociliary clearance prior to infection. Many respiratory viruses bind to glycans on the heavily glycosylated mucins that give mucus its gel-like characteristics. Influenza viruses, some paramyxoviruses, and coronaviruses avoid becoming trapped in the mucus by releasing themselves by means of their envelope-embedded enzymes that destroy glycan receptors. For efficient infection, receptor binding and destruction need to be in balance with the host receptor repertoire. Establishment in a novel host species requires resetting of the balance to adapt to the different glycan repertoire encountered. Growing understanding of species-specific mucosal glycosylation patterns and the dynamic interaction with respiratory viruses identifies the mucus layer as a major host-range determinant and barrier for zoonotic transfer.
Topics: Glycosylation; Host Specificity; Mucins; Mucus; Polysaccharides; Viruses
PubMed: 33875348
DOI: 10.1016/j.tim.2021.03.014 -
Animal : An International Journal of... Mar 2020Making dairy farming more cost-effective and reducing nitrogen environmental pollution could be reached through a reduced input of dietary protein, provided productivity... (Review)
Review
Making dairy farming more cost-effective and reducing nitrogen environmental pollution could be reached through a reduced input of dietary protein, provided productivity is not compromised. This could be achieved through balancing dairy rations for essential amino acids (EAA) rather than their aggregate, the metabolizable protein (MP). This review revisits the estimations of the major true protein secretions in dairy cows, milk protein yield (MPY), metabolic fecal protein (MFP), endogenous urinary loss and scurf and associated AA composition. The combined efficiency with which MP (EffMP) or EAA (EffAA) is used to support protein secretions is calculated as the sum of true protein secretions (MPY + MFP + scurf) divided by the net supply (adjusted to remove the endogenous urinary excretion: MPadj and AAadj). Using the proposed protein and AA secretions, EffMP and EffAA were predicted through meta-analyses (807 treatment means) and validated using an independent database (129 treatment means). The effects of MPadj or AAadj, plus digestible energy intake (DEI), days in milk (DIM) and parity (primiparous v. multiparous), were significant in all models. Models using (MPadj, MPadj × MPadj, DEI and DEI × DEI) or (MPadj/DEI and MPadj/DEI × MPadj/DEI) had similar corrected Akaike's information criterion, but the model using MPadj/DEI performed better in the validation database. A model that also included this ratio was, therefore, used to fitting equations to predict EffAA. These equations predicted well EffAA in the validation database except for Arg which had a strong slope bias. Predictions of MPY from predicted EffMP based on MPadj/DEI, MPadj/DEI × MPadj/DEI, DIM and parity yielded a better fit than direct predictions of MPY based on MPadj, MPadj × MPadj, DEI, DIM and parity. Predictions of MPY based on each EffAA yielded fairly similar results among AA. It is proposed to ponder the mean of MPY predictions obtained from each EffAA by the lowest prediction to retain the potential limitation from AA with the shortest supply. Overall, the revisited estimations of endogenous urinary excretion and MFP, revised AA composition of protein secretions and inclusion of a variable combined EffAA (based on AAadj/DEI, AAadj/DEI × Aadj/DEI, DIM and parity) offer the potential to improve predictions of MPY, identify which AA are potentially in short supply and, therefore, improve the AA balance of dairy rations.
Topics: Amino Acids; Amino Acids, Essential; Animals; Body Weight; Cattle; Diet; Dietary Proteins; Energy Intake; Female; Lactation; Milk; Milk Proteins; Parity; Pregnancy
PubMed: 32024565
DOI: 10.1017/S1751731119003173 -
Frontiers in Immunology 2020Most extracellular proteins are secreted via the classical endoplasmic reticulum (ER)/Golgi-dependent secretion pathway; however, some proteins, including a few... (Review)
Review
Most extracellular proteins are secreted via the classical endoplasmic reticulum (ER)/Golgi-dependent secretion pathway; however, some proteins, including a few danger-associated molecular patterns (DAMPs), are secreted via non-classical ER/Golgi-independent secretion pathways. The evolutionarily conserved high mobility group box1 (HMGB1) is a ubiquitous nuclear protein that can be released by almost all cell types. HMGB1 lacks signal peptide and utilizes diverse non-canonical secretion mechanisms for its extracellular export. Although the post-translational modifications of HMGB1 were demonstrated, the oxidation of HMGB1 and secretion mechanisms are not highlighted yet. We currently investigated that peroxiredoxins I and II (PrxI/II) induce the intramolecular disulfide bond formation of HMGB1 in the nucleus. Disulfide HMGB1 is preferentially transported out of the nucleus by binding to the nuclear exportin chromosome-region maintenance 1 (CRM1). We determined the kinetics of HMGB1 oxidation in bone marrow-derived macrophage as early as a few minutes after lipopolysaccharide treatment, peaking at 4 h while disulfide HMGB1 accumulation was observed within the cells, starting to secrete in the late time point. We have shown that HMGB1 oxidation status, which is known to determine the biological activity in extracellular HMGB1, is crucial for the secretion of HMGB1 from the nucleus. This review summarizes selected aspects of HMGB1 redox biology relevant to the induction and propagation of inflammatory diseases. We implicate the immunological significance and the need for novel HMGB1 inhibitors through mechanism-based studies.
Topics: Animals; HMGB1 Protein; Humans; Oxidation-Reduction; Protein Processing, Post-Translational; Protein Transport
PubMed: 32587593
DOI: 10.3389/fimmu.2020.01189 -
BMC Biology Oct 2023Whereas the translocation of proteins across the cell membrane has been thoroughly investigated, it is still unclear how proteins cross the cell wall in Gram-positive...
BACKGROUND
Whereas the translocation of proteins across the cell membrane has been thoroughly investigated, it is still unclear how proteins cross the cell wall in Gram-positive bacteria, which are widely used for industrial applications. We have studied the secretion of α-amylase AmyE within two different Bacillus strains, B. subtilis and B. licheniformis.
RESULTS
We show that a C-terminal fusion of AmyE with the fluorescent reporter mCherry is secreted via discrete patches showing very low dynamics. These are visible at many places within the cell wall for many minutes. Expression from a high copy number plasmid was required to be able to see these structures we term "secretion zones". Zones corresponded to visualized AmyE activity on the surface of cells, showing that they release active enzymes. They overlapped with SecA signals but did not frequently co-localize with the secretion ATPase. Single particle tracking showed higher dynamics of SecA and of SecDF, involved in AmyE secretion, at the cell membrane than AmyE. These experiments suggest that SecA initially translocates AmyE molecules through the cell membrane, and then diffuses to a different translocon. Single molecule tracking of SecA suggests the existence of three distinct diffusive states of SecA, which change during AmyE overexpression, but increased AmyE secretion does not appear to overwhelm the system.
CONCLUSIONS
Because secretion zones were only found during the transition to and within the stationary phase, diffusion rather than passive transport based on cell wall growth from inside to outside may release AmyE and, thus, probably secreted proteins in general. Our findings suggest active transport through the cell membrane and slow, passive transition through the cell wall, at least for overexpressed proteins, in bacteria of the genus Bacillus.
Topics: Amylases; Bacterial Proteins; Bacillus subtilis; Adenosine Triphosphatases; Protein Transport; Cell Wall; Escherichia coli Proteins
PubMed: 37794427
DOI: 10.1186/s12915-023-01684-1 -
Nucleic Acids Research Jan 2022From industry to food to health, bacteria play an important role in all facets of life. Some of the most important bacteria have been purposely engineered to produce...
From industry to food to health, bacteria play an important role in all facets of life. Some of the most important bacteria have been purposely engineered to produce commercial quantities of antibiotics and therapeutics, and non-classical secretion systems are at the forefront of these technologies. Unlike the classical Sec or Tat pathways, non-classically secreted proteins share few common characteristics and use much more diverse secretion pathways for protein transport. Systematically categorizing and investigating the non-classically secreted proteins will enable a deeper understanding of their associated secretion mechanisms and provide a landscape of the Gram-positive secretion pathway distribution. We therefore developed PncsHub (https://pncshub.erc.monash.edu/), the first universal platform for comprehensively annotating and analyzing Gram-positive bacterial non-classically secreted proteins. PncsHub catalogs 4,914 non-classically secreted proteins, which are delicately categorized into 8 subtypes (including the 'unknown' subtype) and annotated with data compiled from up to 26 resources and visualisation tools. It incorporates state-of-the-art predictors to identify new and homologous non-classically secreted proteins and includes three analytical modules to visualise the relationships between known and putative non-classically secreted proteins. As such, PncsHub aims to provide integrated services for investigating, predicting and identifying non-classically secreted proteins to promote hypothesis-driven laboratory-based experiments.
Topics: Bacterial Proteins; Databases, Protein; Gene Expression Regulation, Bacterial; Gram-Positive Bacteria; Internet; Molecular Sequence Annotation; Phylogeny; Protein Transport; User-Computer Interface
PubMed: 34551435
DOI: 10.1093/nar/gkab814 -
International Journal of Molecular... Nov 2021Most secreted and membrane proteins are targeted to and translocated across the endoplasmic reticulum (ER) membrane through the Sec61 protein-conducting channel.... (Review)
Review
Most secreted and membrane proteins are targeted to and translocated across the endoplasmic reticulum (ER) membrane through the Sec61 protein-conducting channel. Evolutionarily conserved Sec62 and Sec63 associate with the Sec61 channel, forming the Sec complex and mediating translocation of a subset of proteins. For the last three decades, it has been thought that ER protein targeting and translocation occur via two distinct pathways: signal recognition particle (SRP)-dependent co-translational or SRP-independent, Sec62/Sec63 dependent post-translational translocation pathway. However, recent studies have suggested that ER protein targeting and translocation through the Sec translocon are more intricate than previously thought. This review summarizes the current understanding of the molecular functions of Sec62/Sec63 in ER protein translocation.
Topics: Endoplasmic Reticulum; Heat-Shock Proteins; Membrane Transport Proteins; Protein Processing, Post-Translational; Protein Transport; SEC Translocation Channels; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 34884562
DOI: 10.3390/ijms222312757