scholarly journals Therapeutic Nanobodies Targeting Cell Plasma Membrane Transport Proteins: A High-Risk/High-Gain Endeavor

Biomolecules ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 63
Author(s):  
Raf Van Campenhout ◽  
Serge Muyldermans ◽  
Mathieu Vinken ◽  
Nick Devoogdt ◽  
Timo W.M. De Groof
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Membrane Transport ◽  
Transport Proteins ◽  
Therapeutic Drug ◽  
Cell Plasma Membrane ◽  
Plasma Membrane Proteins ◽  
Altered Expression ◽  
Membrane Transport Proteins ◽  
Cell Plasma

Cell plasma membrane proteins are considered as gatekeepers of the cell and play a major role in regulating various processes. Transport proteins constitute a subclass of cell plasma membrane proteins enabling the exchange of molecules and ions between the extracellular environment and the cytosol. A plethora of human pathologies are associated with the altered expression or dysfunction of cell plasma membrane transport proteins, making them interesting therapeutic drug targets. However, the search for therapeutics is challenging, since many drug candidates targeting cell plasma membrane proteins fail in (pre)clinical testing due to inadequate selectivity, specificity, potency or stability. These latter characteristics are met by nanobodies, which potentially renders them eligible therapeutics targeting cell plasma membrane proteins. Therefore, a therapeutic nanobody-based strategy seems a valid approach to target and modulate the activity of cell plasma membrane transport proteins. This review paper focuses on methodologies to generate cell plasma membrane transport protein-targeting nanobodies, and the advantages and pitfalls while generating these small antibody-derivatives, and discusses several therapeutic nanobodies directed towards transmembrane proteins, including channels and pores, adenosine triphosphate-powered pumps and porters.

scholarly journals Membrane proteins synthesized by rabbit reticulocytes.

1975 ◽  
Vol 65 (1) ◽  
pp. 51-64 ◽  
Author(s):  
H F Lodish ◽  
B Small
Keyword(s):  
Amino Acids ◽  
Plasma Membrane ◽  
Membrane Proteins ◽  
Cell Plasma Membrane ◽  
Whole Cells ◽  
Predominant Species ◽  
Cytoplasmic Surface ◽  
Cell Plasma ◽  
Intact Rabbit ◽  
Rabbit Reticulocytes

Intact rabbit reticulocyte cells synthesize two predominant species of polypeptides which are components of the cell plasma membrane. Previous work (Lodish, H. F. 1973. Proc. Natl. Acad. Sci. U. S. A. 70:1526-1530.) showed that these proteins were synthesized by polyribosomes not attached to membranes. We show here that both polypeptides are confined to the cytoplasmic surface of the cell membrane. These studies utilized iodination of whole cells and of membranes with lactoperoxidase, and digestion of whole cells and membranes with chymotrypsin, One of these proteins is synthesized as a precursor, and about 20-40 amino acids are removed after it is incorporated into the membrane, We discuss the probable sites of synthesis of these and other classes of membrane proteins.

scholarly journals PMRT1, a Plasmodium specific parasite plasma membrane transporter is essential for asexual and sexual blood stage development

2021 ◽  
Author(s):  
Jan Stephan Wichers ◽  
Paolo Mesén-Ramírez ◽  
Jing Yu-Strzelczyk ◽  
Gwendolin Fuchs ◽  
Jan Stäcker ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Transport ◽  
Transport Proteins ◽  
Membrane Transporters ◽  
Blood Stage ◽  
Cell Physiology ◽  
Membrane Transporter ◽  
Membrane Hyperpolarization ◽  
Membrane Transport Proteins ◽  
Plasma Membrane Transporter

Membrane transport proteins perform crucial roles in cell physiology. The obligate intracellular parasite Plasmodium falciparum, an agent of human malaria, relies on membrane transport proteins for the uptake of nutrients from the host, disposal of metabolic waste, exchange of metabolites between organelles and generation and maintenance of transmembrane electrochemical gradients for its growth and replication within human erythrocytes. Despite their importance for Plasmodium cellular physiology, the functional roles of a number of membrane transport proteins remain unclear, which is particularly true for orphan membrane transporters that have no or limited sequence homology to transporter proteins in other evolutionary lineages. Therefore, in the current study, we applied endogenous tagging, targeted gene disruption, conditional knockdown and knockout approaches to investigate the subcellular localization and essentiality of six membrane transporters during intraerythrocytic development of P. falciparum parasites. They are localized at different subcellular structures – the food vacuole, the apicoplast, and the parasite plasma membrane – and showed essentiality of four out of the six membrane transporters during asexual development. Additionally, the plasma membrane resident transporter 1 (PMRT1, PF3D7_1135300), a unique Plasmodium-specific plasma membrane transporter, was shown to be essential for gametocytogenesis. Heterologous expression of wild-type and mutation constructs in Xenopus laevis oocytes indicated ion transport upon membrane hyperpolarization and a functional role of negatively charged amino acids protruding into the parasitophorous vacuole lumen. Overall, we reveal the importance of four orphan transporters to blood stage P. falciparum development and provide the first functional characterization of PfPMRT1, an essential parasite membrane transporter.

scholarly journals Loss of the transferrin receptor during the maturation of sheep reticulocytes in vitro. An immunological approach

1983 ◽  
Vol 210 (1) ◽  
pp. 37-47 ◽  
Author(s):  
B T Pan ◽  
R Blostein ◽  
R M Johnstone
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Transferrin Receptor ◽  
Plasma Membranes ◽  
Plasma Membrane Proteins ◽  
Molecular Weights ◽  
Subunit Molecular Weight ◽  
Cell Plasma ◽  
A Subunit

Sheep reticulocyte-specific antiserum absorbed with mature sheep red cells has been used to isolate and identify reticulocyte-specific plasma-membrane proteins and to monitor their loss during incubation in vitro. Specific precipitation of labelled plasma-membrane proteins is obtained when detergent-solubilized extracts of 125I-labelled reticulocyte plasma membranes are incubated with this antiserum and Staphyloccus aureus, but not when mature-cell plasma membranes are treated similarly. During maturation of reticulocytes in vitro (up to 4 days at 37 degrees C), there is a marked decrease in the immunoprecipitable material. The anti-reticulocyte-specific antibodies have been identified as anti-(transferrin receptor) antibodies. By using these antibodies as a probe, the transferrin receptor has been shown to have a subunit molecular weight of 93 000. The data are consistent with reported molecular weights of this receptor and with the proposal that the receptor may exist as a dimer, since [125I]iodotyrosyl-peptide maps of the 93 000- and 186 000-mol.wt. components isolated are shown to be identical. Evidence is presented for the transmembrane nature of the receptor and for the presence of different binding sites for transferrin and these antibodies on the receptor.

scholarly journals Externally disposed plasma membrane proteins. II. Metabolic fate of iodinated polypeptides of mouse L cells.

1975 ◽  
Vol 64 (2) ◽  
pp. 461-479 ◽  
Author(s):  
A L Hubbard ◽  
Z A Cohn
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Sodium Dodecyl ◽  
Relative Distribution ◽  
Plasma Membrane Proteins ◽  
Electron Microscope Autoradiography ◽  
L Cells ◽  
Cell Plasma ◽  
Latex Beads ◽  
Discontinuous Sucrose Gradient

The fate of the L-cell plasma membrane proteins labeled by enzymatic iodination was studied. The disappearance of label from growing cells exhibits a biphasic behavior, with 5-20% lost rapidly (t1/2 similar to 2 h) and 80-90% lost relatively slowly (t1/2 similar to 25-33 h). The loss is temperature dependent and serum independent, and is accompanied by the appearance of 51% (125-I)monoiodotyrosine (MIT) in the medium by 47 h. A variable amount (1-14%) of acid-insoluble label can be recovered in the medium over 47 h. Sodium dodecyl sulfate (SDS)-polyacrylamide gel labeling patterns from cells cultured up to 48 h after iodination reveal no change in the relative distribution of radioactivity, indicating similar rates of degradation for most of the labeled membrane proteins. The fate of the labeled membrane proteins was studied at various times after phagocytosis of nondigestible polystyrene particles. Iodinated L cells phagocytose sufficient 1.1 mum latex beads in 60 min to interiorize 15-30% of the total cell surface area. Electron microscope autoradiography confirmed that labeled membrane is internalized during phagocytosis. The latex-containing phagocytic vacuoles are isolated by flotation in a discontinuous sucrose gradient. 15-30% of the total incorporated label and a comparable percentage of alkaline phosphodiesterase I activity (PDase, a plasma membrane enzyme marker) are recovered in the phagocytic vacuole fraction. Lysosomal enzyme activities are found in the latex vacuole fraction, indicating formation of phagolysosomes. SDS gel analyses reveal that all of the radioactive proteins initially present on the intact cell's surface are interiorized to the same relative extent. Incorporated label and PDase activity disappear much more rapidly from the phagolysosomes than from the whole cell. In the phagolysosomal compartment, greater than 70% of the TCA-precipitable labeled proteins and all of the PDase activity are lost rapidly (t1/2 equals 1-2 h) but similar 30% of the labeled proteins in this compartment are degraded with a 17-20 h half-life. The slowly degraded label is due to specific long-lived polypeptides, of 85,000 and 8,000-15,000 daltons, which remain in the phagolysosomal membrane up to 40 h after phagocytosis.

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