scholarly journals Organization of the gene encoding the human Kell blood group protein [published erratum appears in Blood 1996 Jun 1;87(11):4922]

Blood ◽  
1995 ◽  
Vol 85 (5) ◽  
pp. 1364-1370 ◽  
Author(s):  
S Lee ◽  
E Zambas ◽  
ED Green ◽  
C Redman
Keyword(s):  
Amino Acids ◽  
Blood Group ◽  
Transcription Initiation ◽  
Fetal Liver ◽  
K562 Cells ◽  
Integral Membrane Protein ◽  
Gene Encoding ◽  
Donor And Acceptor ◽  
Exon 1 ◽  
Membrane Spanning

Kell is one of the major blood group systems in human erythrocytes. It is a complex system containing a large number of different antigens. Previously we cloned the Kell cDNA, which was predicted to encode an integral membrane protein with 731 amino acids. Now we have isolated overlapping genomic clones and determined the exon-intron structure of the KEL gene; it spans approximately 21.5 kb with its coding sequence being organized in 19 exons that range in size from 63 bp to 288 bp. The size of introns ranges from 93 bp to approximately 6 kb. The donor and acceptor splice sites all conform to the consensus splicing sequences. Exon 1 encodes only the initiation amino acid, methionine, and contains a consensus Sp1 binding site. The single membrane spanning region of Kell protein is encoded in exon 3 and the putative zinc endopeptidase active site is in exon 16. The amino acids encoded by the 19 exons are identical to those of a person with a common Kell phenotype, as determined by RNA polymerase chain reaction of peripheral blood. Amplification of cDNA 552 ends, derived from human fetal liver, indicated three transcription initiation sites located 30, 81, and 120 bp upstream of the initiation codon. The 552 flanking region of KEL from -176 does not contain a TATA sequence, but has possible GATA-1 binding sites and has significant promoter activity when determined by chloramphenicol acetyltransferase activity in K562 cells.

scholarly journals Characterization of the gene encoding the human LW blood group protein in LW+ and LW- phenotypes

Blood ◽  
1996 ◽  
Vol 87 (7) ◽  
pp. 2962-2967 ◽  
Author(s):  
P Hermand ◽  
PY Le Pennec ◽  
P Rouger ◽  
JP Cartron ◽  
P Bailly
Keyword(s):  
Blood Group ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
K562 Cells ◽  
Gene Encoding ◽  
Potential Binding ◽  
Exon 1 ◽  
Intercellular Adhesion Molecules ◽  
Group Protein

The LW blood group is carried by a 42-kD glycoprotein that belongs to the family of intercellular adhesion molecules. The LW gene is organized into three exons spanning an HindIII fragment of approximately 2.65 kb. The exon/intron architecture correlates to the structural domains of the protein and resembles that of other Ig superfamily members except that the signal peptide and the first Ig- like domain are encoded by the first exon. The 5′UT region (nucleotides -289 to +9) includes potential binding sites for various transcription factors (Ets, CACC, SP1, GATA-1, AP2) and exhibited a significant transcriptional activity after transfection in the erythroleukemic K562 cells. No obvious abnormality of the LW gene, including the 5′UT region, has been detected by sequencing polymerase chain reaction- amplified genomic DNA from RhD+ or RhD- donors and from an Rhnull variant that lacks the Rh and LW proteins on red blood cells. However, a deletion of 10 bp in exon 1 of the LW gene was identified in the genome of an LW (a- b-) individual (Big) deficient for LW antigens but carrying a normal Rh phenotype. The 10-bp deletion generates a premature stop codon and encodes a truncated protein without transmembrane and cytoplasmic domain. No detectable abnormality of the LW gene or transcript could be detected in another LW(a- b-) individual (Nic), suggesting the heterogeneity of these phenotypes.

scholarly journals Transient translocation of the cytoplasmic (endo) domain of a type I membrane glycoprotein into cellular membranes.

1993 ◽  
Vol 120 (4) ◽  
pp. 877-883 ◽  
Author(s):  
N Liu ◽  
D T Brown
Keyword(s):  
Amino Acids ◽  
Membrane Protein ◽  
Signal Sequence ◽  
Attachment Site ◽  
Integral Membrane Protein ◽  
Carboxyl Terminus ◽  
Type I ◽  
E2 Glycoprotein ◽  
Typical Type ◽  
Membrane Spanning

The E2 glycoprotein of the alphavirus Sindbis is a typical type I membrane protein with a single membrane spanning domain and a cytoplasmic tail (endo domain) containing 33 amino acids. The carboxyl terminal domain of the tail has been implicated as (a) attachment site for nucleocapsid protein, and (b) signal sequence for integration of the other alpha-virus membrane proteins 6K and E1. These two functions require that the carboxyl terminus be exposed in the cell cytoplasm (a) and exposed in the lumen of the endoplasmic reticulum (b). We have investigated the orientation of this glycoprotein domain with respect to cell membranes by substituting a tyrosine for the normally occurring serine, four amino acids upstream of the carboxyl terminus. Using radioiodination of this tyrosine as an indication of the exposure of the glycoprotein tail, we have provided evidence that this domain is initially translocated into a membrane and is returned to the cytoplasm after export from the ER. This is the first demonstration of such a transient translocation of a single domain of an integral membrane protein and this rearrangement explains some important aspects of alphavirus assembly.

scholarly journals Lack of the multidrug transporter MRP4/ABCC4 defines the PEL-negative blood group and impairs platelet aggregation

Blood ◽  
2020 ◽  
Vol 135 (6) ◽  
pp. 441-448 ◽  
Author(s):  
Slim Azouzi ◽  
Mahmoud Mikdar ◽  
Patricia Hermand ◽  
Emilie-Fleur Gautier ◽  
Virginie Salnot ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Blood Group ◽  
K562 Cells ◽  
Large Deletion ◽  
Drug Dose ◽  
Compensatory Mechanism ◽  
Loss Of Function ◽  
Gene Encoding ◽  
Whole Exome ◽  
Leukemia Treatment

Abstract The rare PEL-negative phenotype is one of the last blood groups with an unknown genetic basis. By combining whole-exome sequencing and comparative global proteomic investigations, we found a large deletion in the ABCC4/MRP4 gene encoding an ATP-binding cassette (ABC) transporter in PEL-negative individuals. The loss of PEL expression on ABCC4-CRISPR-Cas9 K562 cells and its overexpression in ABCC4-transfected cells provided evidence that ABCC4 is the gene underlying the PEL blood group antigen. Although ABCC4 is an important cyclic nucleotide exporter, red blood cells from ABCC4null/PEL-negative individuals exhibited a normal guanosine 3′,5′-cyclic monophosphate level, suggesting a compensatory mechanism by other erythroid ABC transporters. Interestingly, PEL-negative individuals showed an impaired platelet aggregation, confirming a role for ABCC4 in platelet function. Finally, we showed that loss-of-function mutations in the ABCC4 gene, associated with leukemia outcome, altered the expression of the PEL antigen. In addition to ABCC4 genotyping, PEL phenotyping could open a new way toward drug dose adjustment for leukemia treatment.

scholarly journals AnkB, a Periplasmic Ankyrin-Like Protein inPseudomonas aeruginosa, Is Required for Optimal Catalase B (KatB) Activity and Resistance to Hydrogen Peroxide

2000 ◽  
Vol 182 (16) ◽  
pp. 4545-4556 ◽  
Author(s):  
Michael L. Howell ◽  
Eyad Alsabbagh ◽  
Ju-Fang Ma ◽  
Urs A. Ochsner ◽  
Martin G. Klotz ◽  
...  
Keyword(s):  
Amino Acids ◽  
Hydrogen Peroxide ◽  
Cytoplasmic Membrane ◽  
Membrane Vesicles ◽  
Gene Encoding ◽  
Rnase Protection ◽  
C Terminus ◽  
Hydrophobic Amino Acids ◽  
Increased Sensitivity ◽  
Membrane Spanning

ABSTRACT In this study, we have cloned the ankB gene, encoding an ankyrin-like protein in Pseudomonas aeruginosa. TheankB gene is composed of 549 bp encoding a protein of 183 amino acids that possesses four 33-amino-acid ankyrin repeats that are a hallmark of erythrocyte and brain ankyrins. The location ofankB is 57 bp downstream of katB, encoding a hydrogen peroxide-inducible catalase, KatB. Monomeric AnkB is a 19.4-kDa protein with a pI of 5.5 that possesses 22 primarily hydrophobic amino acids at residues 3 to 25, predicting an inner-membrane-spanning motif with the N terminus in the cytoplasm and the C terminus in the periplasm. Such an orientation in the cytoplasmic membrane and, ultimately, periplasmic space was confirmed using AnkB-BlaM and AnkB-PhoA protein fusions. Circular dichroism analysis of recombinant AnkB minus its signal peptide revealed a secondary structure that is ∼65% α-helical. RNase protection and KatB- and AnkB-LacZ translational fusion analyses indicated that katBand ankB are part of a small operon whose transcription is induced dramatically by H2O2, and controlled by the global transactivator OxyR. Interestingly, unlike the spherical nature of ankyrin-deficient erythrocytes, the cellular morphology of anankB mutant was identical to that of wild-type bacteria, yet the mutant produced more membrane vesicles. The mutant also exhibited a fourfold reduction in KatB activity and increased sensitivity to H2O2, phenotypes that could be complemented in trans by a plasmid constitutively expressing ankB. Our results suggest that AnkB may form an antioxidant scaffolding with KatB in the periplasm at the cytoplasmic membrane, thus providing a protective lattice work for optimal H2O2 detoxification.

scholarly journals cDNA cloning, genomic organization and chromosomal localization of human heparan glucosaminyl N-deacetylase/N-sulphotransferase-2

1998 ◽  
Vol 332 (2) ◽  
pp. 303-307 ◽  
Author(s):  
Donald E. HUMPHRIES ◽  
Julia LANCIOTTI ◽  
Joel B. KARLINSKY
Keyword(s):  
Amino Acids ◽  
Chromosomal Localization ◽  
Amino Acid Sequences ◽  
Gene Encoding ◽  
Exon 2 ◽  
Ancestral Gene ◽  
Exon 1 ◽  
Transmembrane Regions ◽  
Conserved Amino Acids

The cDNA and gene encoding human heparan glucosaminyl N-deacetylase/N-sulphotransferase-2 have been cloned. The cDNA encoded a protein of 883 amino acids that was 94% similar to heparan N-sulphotransferase-2 from mouse mast cells. Comparison of the deduced amino acid sequences of human heparan N-sulphotransferase-1 and -2 showed that the enzymes were 70% similar; greater than 90% of the amino acids between residues 418 and 543 were identical. The least conserved amino acids were found in the N-terminus/putative transmembrane regions of the two enzymes. The human heparan N-sulphotransferase-2 gene was localized to chromosome arm 10q (band 10q22) by in situ fluorescent hybridization. The gene contains 13 exons spanning 6.5 kb, ranging in size from 88 bp (exon 2) to > 1 kb (exon 1), and 12 introns, which were found to occur at similar sites within the coding sequence of the human heparan N-sulphotransferase-1 gene. The structure of the two genes differed in that the heparan N-sulphotransferase-1 gene contained one additional intron. The similarity of the heparan N-sulphotransferase-1 and -2 proteins and their similar exon-intron organization suggest that they derive from a common ancestral gene.

scholarly journals Identification of a novel homozygous loss-of-function mutation in FUCA1 gene causing severe fucosidosis: A case report

2021 ◽  
Vol 49 (4) ◽  
pp. 030006052110059
Author(s):  
Xinwen Zhang ◽  
Shaozhi Zhao ◽  
Hongwei Liu ◽  
Xiaoyan Wang ◽  
Xiaolei Wang ◽  
...  
Keyword(s):  
Amino Acids ◽  
Lysosomal Storage Disorder ◽  
Autosomal Recessive ◽  
Signs And Symptoms ◽  
Lysosomal Storage ◽  
Loss Of Function ◽  
Wild Type ◽  
Single Nucleotide ◽  
Whole Exome ◽  
Exon 1

Fucosidosis is a rare lysosomal storage disorder characterized by deficiency of α-L-fucosidase with an autosomal recessive mode of inheritance. Here, we describe a 4-year-old Chinese boy with signs and symptoms of fucosidosis but his parents were phenotypically normal. Whole exome sequencing (WES) identified a novel homozygous single nucleotide deletion (c.82delG) in the exon 1 of the FUCA1 gene. This mutation will lead to a frameshift which will result in the formation of a truncated FUCA1 protein (p.Val28Cysfs*105) of 132 amino acids approximately one-third the size of the wild type FUCA1 protein (466 amino acids). Both parents were carrying the mutation in a heterozygous state. This study expands the mutational spectrum of the FUCA1 gene associated with fucosidosis and emphasises the benefits of WES for accurate and timely clinical diagnosis of this rare disease.

scholarly journals GIT1, a Gene Encoding a Novel Transporter for Glycerophosphoinositol in Saccharomyces cerevisiae

Genetics ◽  
1998 ◽  
Vol 149 (4) ◽  
pp. 1707-1715 ◽  
Author(s):  
J L Patton-Vogt ◽  
S A Henry
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Regulatory Gene ◽  
Sugar Transport ◽  
Open Reading Frame ◽  
Gene Encoding ◽  
Reading Frame ◽  
Membrane Spanning ◽  
Membrane Spanning Domains ◽  
Uptake And Metabolism ◽  
Cells Cultured

Abstract Phosphatidylinositol catabolism in Saccharomyces cerevisiae cells cultured in media containing inositol results in the release of glycerophosphoinositol (GroPIns) into the medium. As the extracellular concentration of inositol decreases with growth, the released GroPIns is transported back into the cell. Exploiting the ability of the inositol auxotroph, ino1, to use exogenous GroPIns as an inositol source, we have isolated mutants (Git−) defective in the uptake and metabolism of GroPIns. One mutant was found to be affected in the gene encoding the transcription factor, SPT7. Mutants of the positive regulatory gene INO2, but not of its partner, INO4, also have the Git− phenotype. Another mutant was complemented by a single open reading frame (ORF) termed GIT1 (glycerophosphoinositol). This ORF consists of 1556 bp predicted to encode a polypeptide of 518 amino acids and 57.3 kD. The predicted Git1p has similarity to a variety of S. cerevisiae transporters, including a phosphate transporter (Pho84p), and both inositol transporters (Itr1p and Itr2p). Furthermore, Git1p contains a sugar transport motif and 12 potential membrane-spanning domains. Transport assays performed on a git1 mutant together with the above evidence indicate that the GIT1 gene encodes a permease involved in the uptake of GroPIns.

SPT15, the gene encoding the yeast TATA binding factor TFIID, is required for normal transcription initiation in vivo

Cell ◽  
1989 ◽  
Vol 58 (6) ◽  
pp. 1183-1191 ◽  
Author(s):  
David M. Eisenmann ◽  
Catherine Dollard ◽  
Fred Winston
Keyword(s):  
Transcription Initiation ◽  
Gene Encoding ◽  
Binding Factor

Active amino acids of the Kell blood group protein and model of the ectodomain based on the structure of neutral endopeptidase 24.11

Blood ◽  
2003 ◽  
Vol 102 (8) ◽  
pp. 3028-3034 ◽  
Author(s):  
Soohee Lee ◽  
Asim K. Debnath ◽  
Colvin M. Redman
Keyword(s):  
Amino Acids ◽  
Blood Group ◽  
Active Site ◽  
Neutral Endopeptidase ◽  
Site Directed Mutagenesis ◽  
Peptide Hydrolysis ◽  
Enzyme Active Site ◽  
Endopeptidase 24.11 ◽  
Outer Domain ◽  
Group Protein

Abstract In addition to its importance in transfusion, Kell protein is a member of the M13 family of zinc endopeptidases and functions as an endothelin-3–converting enzyme. To obtain information on the structure of Kell protein we built a model based on the crystal structure of the ectodomain of neutral endopeptidase 24.11 (NEP). Similar to NEP, the Kell protein has 2 globular domains consisting mostly of α-helical segments. The domain situated closest to the membrane contains both the N- and C-terminal sequences and the enzyme-active site. The outer domain contains all of the amino acids whose substitutions lead to different Kell blood group phenotypes. In the model, the zinc peptidase inhibitor, phosphoramidon, was docked in the active site. Site-directed mutagenesis of amino acids in the active site was performed and the enzymatic activities of expressed mutant Kell proteins analyzed and compared with NEP. Our studies indicate that Kell and NEP use the same homologous amino acids in the coordination of zinc and in peptide hydrolysis. However, Kell uses different amino acids than NEP in substrate binding and appears to have more flexibility in the composition of amino acids allowed in the active site.

scholarly journals The Pichia pastoris PER6 gene product is a peroxisomal integral membrane protein essential for peroxisome biogenesis and has sequence similarity to the Zellweger syndrome protein PAF-1.

1996 ◽  
Vol 16 (5) ◽  
pp. 2527-2536 ◽  
Author(s):  
H R Waterham ◽  
Y de Vries ◽  
K A Russel ◽  
W Xie ◽  
M Veenhuis ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Membrane Protein ◽  
Sequence Similarity ◽  
Zellweger Syndrome ◽  
Methylotrophic Yeast ◽  
Integral Membrane Protein ◽  
Biochemical Properties ◽  
Podospora Anserina ◽  
Peroxisome Biogenesis ◽  
Membrane Spanning

We report the cloning of PER6, a gene essential for peroxisome biogenesis in the methylotrophic yeast Pichia pastoris. The PER6 sequence predicts that its product Per6p is a 52-kDa polypeptide with the cysteine-rich C3HC4 motif. Per6p has significant overall sequence similarity with the human peroxisome assembly factor PAF-1, a protein that is defective in certain patients suffering from the peroxisomal disorder Zellweger syndrome, and with car1, a protein required for peroxisome biogenesis and caryogamy in the filamentous fungus Podospora anserina. In addition, the C3HC4 motif and two of the three membrane-spanning segments predicted for Per6p align with the C3HC4 motifs and the two membrane-spanning segments predicted for PAF-1 and car1. Like PAF-1, Per6p is a peroxisomal integral membrane protein. In methanol- or oleic acid-induced cells of per6 mutants, morphologically recognizable peroxisomes are absent. Instead, peroxisomal remnants are observed. In addition, peroxisomal matrix proteins are synthesized but located in the cytosol. The similarities between Per6p and PAF-1 in amino acid sequence and biochemical properties, and between mutants defective in their respective genes, suggest that Per6p is the putative yeast homolog of PAF-1.

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