Different molecular forms of rat kidney gp330, the dominant autoantigen of active Heymann nephritis

The primary structure, consisting of 1650 amino acid residues, of the C-terminal end of the dominant autoantigen of active Heymann Nephritis, gp330, from rat kidney was obtained by cloning and sequencing of cDNA clones. Comparison of this sequence with the previously published sequences of fragments of the C-terminal end of gp330 [Raychowdhury, Niles, McCluskey and Smith (1989) Science 244, 1163-1165] revealed certain differences in their primary structures. These differences included several single amino acid substitutions, replacement of a stretch of 15 amino acid residues by a different stretch of six amino acid residues, and different lengths of cytoplasmic domain (188 versus 213 amino acid residues). These findings of two different primary structures of gp330 provide direct evidence for the existence of two molecular forms of gp330.

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Molecular and Functional Analyses of Kunjin Virus Infectious cDNA Clones Demonstrate the Essential Roles for NS2A in Virus Assembly and for a Nonconservative Residue in NS3 in RNA Replication

Journal of Virology ◽

10.1128/jvi.77.14.7804-7813.2003 ◽

2003 ◽

Vol 77 (14) ◽

pp. 7804-7813 ◽

Cited By ~ 123

Author(s):

Wen Jun Liu ◽

Hua Bo Chen ◽

Alexander A. Khromykh

Keyword(s):

Amino Acid ◽

Virus Assembly ◽

Nonstructural Protein ◽

Rna Replication ◽

Site Directed Mutagenesis ◽

Single Amino Acid ◽

Cdna Clones ◽

Amino Acid Residues ◽

Wild Type ◽

Common Amino Acid

ABSTRACT A number of full-length cDNA clones of Kunjin virus (KUN) were previously prepared; it was shown that two of them, pAKUN and FLSDX, differed in specific infectivities of corresponding in vitro transcribed RNAs by ∼100,000-fold (A. A. Khromykh et al., J. Virol. 72:7270-7279, 1998). In this study, we analyzed a possible genetic determinant(s) of the observed differences in infectivity initially by sequencing the entire cDNAs of both clones and comparing them with the published sequence of the parental KUN strain MRM61C. We found six common amino acid residues in both cDNA clones that were different from those in the published MRM61C sequence but were similar to those in the published sequences of other flaviviruses from the same subgroup. pAKUN clone had four additional codon changes, i.e., Ile59 to Asn and Arg175 to Lys in NS2A and Tyr518 to His and Ser557 to Pro in NS3. Three of these substitutions except the previously shown marker mutation, Arg175 to Lys in NS2A, reverted to the wild-type sequence in the virus eventually recovered from pAKUN RNA-transfected BHK cells, demonstrating the functional importance of these residues in viral replication and/or viral assembly. Exchange of corresponding DNA fragments between pAKUN and FLSDX clones and site-directed mutagenesis revealed that the Tyr518-to-His mutation in NS3 was responsible for an ∼5-fold decrease in specific infectivity of transcribed RNA, while the Ile59-to-Asn mutation in NS2A completely blocked virus production. Correction of the Asn59 in pAKUN NS2A to the wild-type Ile residue resulted in complete restoration of RNA infectivity. Replication of KUN replicon RNA with an Ile59-to-Asn substitution in NS2A and with a Ser557-to-Pro substitution in NS3 was not affected, while the Tyr518-to-His substitution in NS3 led to severe inhibition of RNA replication. The impaired function of the mutated NS2A in production of infectious virus was complemented in trans by the helper wild-type NS2A produced from the KUN replicon RNA. However, replicon RNA with mutated NS2A could not be packaged in trans by the KUN structural proteins. The data demonstrated essential roles for the KUN nonstructural protein NS2A in virus assembly and for NS3 in RNA replication and identified specific single-amino-acid residues involved in these functions.

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Cleavage of single amino acid residues from Merrifield resin with hydrogen chloride and hydrogen fluoride

The Journal of Organic Chemistry ◽

10.1021/jo00834a067 ◽

1970 ◽

Vol 35 (9) ◽

pp. 3151-3152 ◽

Cited By ~ 67

Author(s):

J. Scotchler ◽

R. Lozier ◽

Arthur Brouhard. Robinson

Keyword(s):

Amino Acid ◽

Hydrogen Chloride ◽

Hydrogen Fluoride ◽

Single Amino Acid ◽

Amino Acid Residues ◽

Merrifield Resin

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Molecular cloning of two human liver 3 α-hydroxysteroid/dihydrodiol dehydrogenase isoenzymes that are identical with chlordecone reductase and bile-acid binder

Biochemical Journal ◽

10.1042/bj2990545 ◽

1994 ◽

Vol 299 (2) ◽

pp. 545-552 ◽

Cited By ~ 63

Author(s):

Y Deyashiki ◽

A Ogasawara ◽

T Nakayama ◽

M Nakanishi ◽

Y Miyabe ◽

...

Keyword(s):

Bile Acid ◽

Amino Acid ◽

Human Liver ◽

Polyclonal Antibodies ◽

Amino Acid Sequences ◽

Cdna Clones ◽

Amino Acid Residues ◽

Human Bile ◽

Coding Regions ◽

Computer Based

Human liver contains two dihydrodiol dehydrogenases, DD2 and DD4, associated with 3 alpha-hydroxysteroid dehydrogenase activity. We have raised polyclonal antibodies that cross-reacted with the two enzymes and isolated two 1.2 kb cDNA clones (C9 and C11) for the two enzymes from a human liver cDNA library using the antibodies. The clones of C9 and C11 contained coding sequences corresponding to 306 and 321 amino acid residues respectively, but lacked 5′-coding regions around the initiation codon. Sequence analyses of several peptides obtained by enzymic and chemical cleavages of the two purified enzymes verified that the C9 and C11 clones encoded DD2 and DD4 respectively, and further indicated that the sequence of DD2 had at least additional 16 residues upward from the N-terminal sequence deduced from the cDNA. There was 82% amino acid sequence identity between the two enzymes, indicating that the enzymes are genetic isoenzymes. A computer-based comparison of the cDNAs of the isoenzymes with the DNA sequence database revealed that the nucleotide and amino acid sequences of DD2 and DD4 are virtually identical with those of human bile-acid binder and human chlordecone reductase cDNAs respectively.

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Identification of critical amino acid residues in the regulatory N-terminal domain of PMEL

Scientific Reports ◽

10.1038/s41598-021-87259-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Susan M. Mitchell ◽

Morven Graham ◽

Xinran Liu ◽

Ralf M. Leonhardt

Keyword(s):

Amino Acid ◽

Pigment Cell ◽

Specific Protein ◽

Single Amino Acid ◽

Amyloid Formation ◽

Amino Acid Residues ◽

Proteolytic Fragment ◽

The Core ◽

N Terminus ◽

In Cis

AbstractThe pigment cell-specific protein PMEL forms a functional amyloid matrix in melanosomes onto which the pigment melanin is deposited. The amyloid core consists of a short proteolytic fragment, which we have termed the core-amyloid fragment (CAF) and perhaps additional parts of the protein, such as the PKD domain. A highly O-glycosylated repeat (RPT) domain also derived from PMEL proteolysis associates with the amyloid and is necessary to establish the sheet-like morphology of the assemblies. Excluded from the aggregate is the regulatory N-terminus, which nevertheless must be linked in cis to the CAF in order to drive amyloid formation. The domain is then likely cleaved away immediately before, during, or immediately after the incorporation of a new CAF subunit into the nascent amyloid. We had previously identified a 21 amino acid long region, which mediates the regulatory activity of the N-terminus towards the CAF. However, many mutations in the respective segment caused misfolding and/or blocked PMEL export from the endoplasmic reticulum, leaving their phenotype hard to interpret. Here, we employ a saturating mutagenesis approach targeting the motif at single amino acid resolution. Our results confirm the critical nature of the PMEL N-terminal region and identify several residues essential for PMEL amyloidogenesis.

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A Mutation in the  Subunit of the Platelet Integrin IIbβ3 Identifies a Novel Region Important for Ligand Binding

Blood ◽

10.1182/blood.v93.3.918 ◽

1999 ◽

Vol 93 (3) ◽

pp. 918-924 ◽

Cited By ~ 19

Author(s):

Eileen Collins Tozer ◽

Elizabeth K. Baker ◽

Mark H. Ginsberg ◽

Joseph C. Loftus

Keyword(s):

Amino Acid ◽

Ligand Binding ◽

Single Amino Acid ◽

Chimeric Receptor ◽

Amino Acid Residues ◽

Genetic Approach ◽

Specific Amino Acid ◽

Transfected Cells ◽

Binding Function ◽

A Cell

Abstract An unbiased genetic approach was used to identify a specific amino acid residue in the IIb subunit important for the ligand binding function of the integrin IIbβ. Chemically mutagenized cells were selected by flow cytometry based on their inability to bind the ligand mimetic antibody PAC1 and a cell line containing a single amino acid substitution in IIb at position 224 (D→V) was identified. Although well expressed on the surface of transfected cells, IIbD224Vβ3 as well as IIbD224Aβ3 did not bind IIbβ3-specific ligands or a RGD peptide, a ligand shared in common with vβ3. Insertion of exon 5 of IIb, residues G193-W235, into the backbone of the v subunit did not enable the chimeric receptor to bind IIbβ3-specific ligands. However, the chimeric receptor was still capable of binding to a RGD affinity matrix. IIbD224 is not well conserved among other integrin  subunits and is located in a region of significant variability. In addition, amino acid D224 lies within a predicted loop of the recently proposed β-propeller model for integrin  subunits and is adjacent to a loop containing amino acid residues previously implicated in receptor function. These data support a role for this region in ligand binding function of the IIbβ3 receptor.

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Cloning and sequencing of protochlorophyllide reductase

Biochemical Journal ◽

10.1042/bj2650789 ◽

1990 ◽

Vol 265 (3) ◽

pp. 789-798 ◽

Cited By ~ 46

Author(s):

P M Darrah ◽

S A Kay ◽

G R Teakle ◽

W T Griffiths

Keyword(s):

Amino Acid ◽

Chemical Analysis ◽

Amino Acid Sequence ◽

Cdna Library ◽

Avena Sativa ◽

Cdna Clones ◽

Lambda Phage ◽

Cloning And Sequencing ◽

Protochlorophyllide Reductase ◽

Cnbr Cleavage

Putative protochlorophyllide reductase cDNA clones (252 and 113) were isolated from an etiolated-oat (Avena sativa) cDNA library. These were used to indirectly characterize a further clone, p127, isolated from a lambda-phage gt11 cDNA library. The latter (1.15 kb in length) was sequenced, and the derived amino acid sequence was shown to be remarkably similar to that derived from chemical analysis of a CNBr-cleavage fragment of the purified reductase, p127 codes for more than 95% of the reductase protein.

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Single Amino Acid Residues in the E- and P-selectin Epidermal Growth Factor Domains Can Determine Carbohydrate Binding Specificity

Journal of Biological Chemistry ◽

10.1074/jbc.271.27.16160 ◽

1996 ◽

Vol 271 (27) ◽

pp. 16160-16170 ◽

Cited By ~ 79

Author(s):

B. Mitch Revelle ◽

Dee Scott ◽

Pamela J. Beck

Keyword(s):

Amino Acid ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Binding Specificity ◽

Single Amino Acid ◽

Carbohydrate Binding ◽

Amino Acid Residues ◽

Epidermal Growth

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Amino Acid Residues in the Carboxy-Terminal Region of Cottontail Rabbit Papillomavirus E6 Influence Spontaneous Regression of Cutaneous Papillomas

Journal of Virology ◽

10.1128/jvi.76.23.11801-11808.2002 ◽

2002 ◽

Vol 76 (23) ◽

pp. 11801-11808 ◽

Cited By ~ 27

Author(s):

Jiafen Hu ◽

Nancy M. Cladel ◽

Martin D. Pickel ◽

Neil D. Christensen

Keyword(s):

Amino Acid ◽

Immune Responses ◽

Single Amino Acid ◽

Human Populations ◽

Amino Acid Residues ◽

High Frequencies ◽

Host Immune Responses ◽

Cottontail Rabbit ◽

Single Amino Acid Change ◽

Carboxy Terminal

ABSTRACT Previous studies have identified two different strains of cottontail rabbit papillomavirus (CRPV) that differ by approximately 5% in base pair sequence and that perform quite differently when used to challenge New Zealand White (NZW) rabbit skin. One strain caused persistent lesions (progressor strain), and the other induced papillomas that spontaneously regressed (regressor strain) at high frequencies (J. Salmon, M. Nonnenmacher, S. Caze, P. Flamant, O. Croissant, G. Orth, and F. Breitburd, J. Virol. 74:10766-10777, 2000; J. Salmon, N. Ramoz, P. Cassonnet, G. Orth, and F. Breitburd, Virology 235:228-234, 1997). We generated a panel of CRPV genomes that contained chimeric and mutant progressor and regressor strain E6 genes and assessed the outcome upon infection of both outbred and EIII/JC inbred NZW rabbits. The carboxy-terminal 77-amino-acid region of the regressor CRPV strain E6, which contained 15 amino acid residues that are different from those of the equivalent region of the persistent CRPV strain E6, played a dominant role in the conversion of the persistent CRPV strain to one showing high rates of spontaneous regressions. In addition, a single amino acid change (G252E) in the E6 protein of the CRPV progressor strain led to high frequencies of spontaneous regressions in inbred rabbits. These observations imply that small changes in the amino acid sequences of papillomavirus proteins can dramatically impact the outcome of natural host immune responses to these viral infections. The data imply that intrastrain differences between separate isolates of a single papillomavirus type (such as human papillomavirus type 16) may contribute to a collective variability in host immune responses in outbred human populations.

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Amino acid sequence of band-3 protein from rainbow trout erythrocytes derived from cDNA

Biochemical Journal ◽

10.1042/bj2850017 ◽

1992 ◽

Vol 285 (1) ◽

pp. 17-23 ◽

Cited By ~ 46

Author(s):

S Hübner ◽

F Michel ◽

V Rudloff ◽

H Appelhans

Keyword(s):

Rainbow Trout ◽

Amino Acid ◽

Amino Acid Sequence ◽

Untranslated Region ◽

Band 3 ◽

Cdna Clones ◽

Amino Acid Residues ◽

Band 3 Protein ◽

Lower Vertebrate ◽

Calculated Molecular Mass

In this report we present the first complete band-3 cDNA sequence of a poikilothermic lower vertebrate. The primary structure of the anion-exchange protein band 3 (AE1) from rainbow trout erythrocytes was determined by nucleotide sequencing of cDNA clones. The overlapping clones have a total length of 3827 bp with a 5′-terminal untranslated region of 150 bp, a 2754 bp open reading frame and a 3′-untranslated region of 924 bp. Band-3 protein from trout erythrocytes consists of 918 amino acid residues with a calculated molecular mass of 101 827 Da. Comparison of its amino acid sequence revealed a 60-65% identity within the transmembrane spanning sequence of band-3 proteins published so far. An additional insertion of 24 amino acid residues within the membrane-associated domain of trout band-3 protein was identified, which until now was thought to be a general feature only of mammalian band-3-related proteins.

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