Identification, purification, and partial characterization of a novel Mr 28,000 integral membrane protein from erythrocytes and renal tubules.

A monoclonal antibody (MAb 30B6) was recently described by Rogalski and Singer (J. Cell Biol. 101:785-801, 1985) which identified an integral membrane glycoprotein of chicken cells that was associated with a wide variety of sites of actin microfilament attachments to membranes. In this report, we present a further characterization of this integral protein. An immunochemical comparison was made of MAb 30B6 binding properties with those of two other MAbs, JG9 and JG22, which identify a component of a membrane protein complex that interacts with extracellular matrix proteins including fibronectin. We showed that the 110-kilodalton protein recognized by MAb 30B6 in extracts of chicken gizzard smooth muscle is identical, or closely related, to the protein that reacts with MAbs JG9 and JG22. These 110-kilodalton proteins are also structurally closely similar, if not identical, to one another as demonstrated by 125I-tryptic peptide maps. However, competition experiments showed that MAb 30B6 recognizes a different epitope from those recognized by MAbs JG9 and JG22. In addition, the 30B6 antigen is part of a complex that can be isolated on fibronectin columns. These results together establish that the 30B6 antigen is the same as, or closely similar to, the beta-chain of the protein complex named integrin, which is the complex on chicken fibroblast membranes that binds fibronectin. Although the 30B6 antigen is present in a wide range of tissues, its apparent molecular weight on gels varies in different tissues. These differences in apparent molecular weight are due, in large part, to differences in glycosylation.

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Characterization of an 80-kDa phosphoprotein involved in parietal cell stimulation

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1989.256.6.g1070 ◽

1989 ◽

Vol 256 (6) ◽

pp. G1070-G1081 ◽

Cited By ~ 13

Author(s):

T. Urushidani ◽

D. K. Hanzel ◽

J. G. Forte

Keyword(s):

Membrane Protein ◽

Parietal Cell ◽

Apical Membrane ◽

Peptide Mapping ◽

Integral Membrane Protein ◽

Gastric Glands ◽

Low Speed ◽

Calcium Dependent ◽

Apical Membranes

When isolated rabbit gastric glands were stimulated with histamine plus isobutylmethylxanthine, a redistribution of H+-K+-ATPase, from microsomes to a low-speed pellet, occurred in association with the phosphorylation of an 80-kDa protein (80K) in the apical membrane-rich fraction purified from the low-speed pellet. Histamine alone or dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP), but not carbachol, also stimulated both the redistribution of H+-K+-ATPase and phosphorylation of 80K. Under stimulated conditions, 80K copurified in the apical membrane fraction along with H+-K+-ATPase and actin; whereas purified microsomes from resting stomach were highly enriched in H+-K+-ATPase but contained neither 80K nor actin. Treatment of the apical membranes with detergents, salts, sonication, and so on, led us to conclude that 80K is a membrane protein, unlike actin; however, the mode of association of 80K with membrane differed from H+-K+-ATPase, an integral membrane protein. Isoelectric focusing and peptide mapping revealed that 80K consists of six isomers of slightly differing pI, with 32P occurring only in the three most acidic isomers and exclusively on serine residues. Moreover, stimulation elicited a shift in the amount of 80K isomers, from basic to acidic, as well as phosphorylation. We conclude that 80K is an apical membrane protein in the parietal cell and an important substrate for cAMP-dependent, but not calcium-dependent, pathway of acid secretion.

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Cloning and characterization of the SmIMP25 integral membrane protein of the parasitic helminth Schistosoma mansoni

Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression ◽

10.1016/0167-4781(94)90178-3 ◽

1994 ◽

Vol 1218 (3) ◽

pp. 273-282 ◽

Cited By ~ 6

Author(s):

Alexander Markovics ◽

Daniela Ram ◽

Grossman Zehava ◽

Etty Ziv ◽

Frida Lantner ◽

...

Keyword(s):

Membrane Protein ◽

Schistosoma Mansoni ◽

Integral Membrane Protein ◽

Parasitic Helminth

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Characterization of YmgF, a 72-Residue Inner Membrane Protein That Associates with the Escherichia coli Cell Division Machinery

Journal of Bacteriology ◽

10.1128/jb.00331-08 ◽

2008 ◽

Vol 191 (1) ◽

pp. 333-346 ◽

Cited By ~ 44

Author(s):

Gouzel Karimova ◽

Carine Robichon ◽

Daniel Ladant

Keyword(s):

Escherichia Coli ◽

Cell Division ◽

Membrane Protein ◽

Integral Membrane Protein ◽

Dependent Manner ◽

E Coli ◽

Division Site ◽

Inner Membrane Protein ◽

Two Hybrid

ABSTRACT Formation of the Escherichia coli division septum is catalyzed by a number of essential proteins (named Fts) that assemble into a ring-like structure at the future division site. Many of these Fts proteins are intrinsic transmembrane proteins whose functions are largely unknown. In the present study, we attempted to identify a novel putative component(s) of the E. coli cell division machinery by searching for proteins that could interact with known Fts proteins. To do that, we used a bacterial two-hybrid system based on interaction-mediated reconstitution of a cyclic AMP (cAMP) signaling cascade to perform a library screening in order to find putative partners of E. coli cell division protein FtsL. Here we report the characterization of YmgF, a 72-residue integral membrane protein of unknown function that was found to associate with many E. coli cell division proteins and to localize to the E. coli division septum in an FtsZ-, FtsA-, FtsQ-, and FtsN-dependent manner. Although YmgF was previously shown to be not essential for cell viability, we found that when overexpressed, YmgF was able to overcome the thermosensitive phenotype of the ftsQ1(Ts) mutation and restore its viability under low-osmolarity conditions. Our results suggest that YmgF might be a novel component of the E. coli cell division machinery.

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Isolation and partial characterization of antibody- and globin-enriched complexes from membranes of dense human erythrocytes

Biochemical Journal ◽

10.1042/bj2780057 ◽

1991 ◽

Vol 278 (1) ◽

pp. 57-62 ◽

Cited By ~ 46

Author(s):

R Kannan ◽

J Yuan ◽

P S Low

Keyword(s):

Membrane Protein ◽

Cell Surface ◽

Integral Membrane Protein ◽

Protein Band ◽

Reticuloendothelial System ◽

Band 3 ◽

Isolation And Characterization ◽

Protein Clusters ◽

Associated Proteins

In previous studies we have described a process whereby an erythrocyte in biochemical distress can initiate its own removal by macrophages of the reticuloendothelial system. This process involves the clustering of the integral membrane protein band 3 by denatured haemoglobin and the subsequent recognition of the exofacial poles of clustered band 3 and associated proteins by autologous antibodies. To determine whether this clearance pathway might mediate normal cell turnover, the fraction of normal erythrocytes containing the 0.5% densest cells, which are known to be destined for immediate removal, was isolated and characterized biochemically. This densest fraction was found to contain 6 times more membrane-bound globin (haemichromes) and 10 times more surface-bound autologous IgG than the other fractions containing cells of lower density. To determine whether the autologous IgG was physically associated with the haemichrome-stabilized membrane protein clusters, a procedure was developed for isolation and characterization of the microscopic aggregates. The isolated aggregates were found to contain a disulphide-cross-linked mixture of several membrane proteins, predominantly haemichromes, spectrin and band 3. Although the aggregates constituted only 0.09% of the total membrane protein, they still contained approximately 55% of the total cell-surface IgG. Since in control studies anti-(blood group A) antibodies, which are distributed randomly over the surface of type A cells, could not be recovered in the aggregate, we conclude that the autologous cell-surface IgGs were physically associated with the membrane protein clusters when they were co-isolated with them in our procedure. Thus the 640-fold enrichment of autologous IgG in the aggregates compared with regions of the membrane devoid of tightly clustered protein suggests that sites of integral protein clustering either are non-specifically sticky to IgG or are viewed as foreign or ‘non-self’ by the immune system and aggressively opsonized with IgG.

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The Hansenula polymorpha PER8 gene encodes a novel peroxisomal integral membrane protein involved in proliferation.

The Journal of Cell Biology ◽

10.1083/jcb.128.3.307 ◽

1995 ◽

Vol 128 (3) ◽

pp. 307-319 ◽

Cited By ~ 90

Author(s):

X Tan ◽

H R Waterham ◽

M Veenhuis ◽

J M Cregg

Keyword(s):

Amino Acid ◽

Membrane Protein ◽

Hansenula Polymorpha ◽

Methylotrophic Yeast ◽

Integral Membrane Protein ◽

Peroxisome Biogenesis ◽

Primary Sequence ◽

Zinc Finger Motifs ◽

Molecular Machinery

We previously described the isolation of mutants of the methylotrophic yeast Hansenula polymorpha that are defective in peroxisome biogenesis. Here, we describe the characterization of one of these mutants, per8, and the cloning of the PER8 gene. In either methanol or methylamine medium, conditions that normally induce the organelles, per8 cells contain no peroxisome-like structures and peroxisomal enzymes are located in the cytosol. The sequence of PER8 predicts that its product (Per8p) is a novel polypeptide of 34 kD, and antibodies against Per8p recognize a protein of 31 kD. Analysis of the primary sequence of Per8p revealed a 39-amino-acid cysteine-rich segment with similarity to the C3HC4 family of zinc-finger motifs. Overexpression of PER8 results in a markedly enhanced increase in peroxisome numbers. We show that Per8p is an integral membrane protein of the peroxisome and that it is concentrated in the membranes of newly formed organelles. We propose that Per8p is a component of the molecular machinery that controls the proliferation of this organelle.

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