scholarly journals Monoclonal antibodies to CD44 and their influence on hyaluronan recognition.

1995 ◽  
Vol 130 (2) ◽  
pp. 485-495 ◽  
Author(s):  
Z Zheng ◽  
S Katoh ◽  
Q He ◽  
K Oritani ◽  
K Miyake ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Amino Acid ◽  
Tumor Metastasis ◽  
Single Amino Acid ◽  
Cross Linking ◽  
Amino Acid Substitutions ◽  
Antibody Specificity ◽  
Lymphocyte Migration ◽  
Antibody Recognition ◽  
Blocking Antibodies

Antibodies to CD44 have been used to inhibit a variety of processes which include lymphohemopoiesis, lymphocyte migration, and tumor metastasis. Some, but not all, CD44-mediated functions derive from its ability to serve as a receptor for hyaluronan (HA). However, sites on CD44 that interact with either ligands or antibodies are poorly understood. Interspecies rat/mouse CD44 chimeras were used to analyze the specificity of 25 mAbs and to determine that they recognize at least seven epitopes. Amino acid substitutions that resulted in loss of antibody recognition were all located in the region of homology to other cartilage link family proteins. While at least five epitopes were eliminated by single amino acid replacements, multiple residues had to be changed to destroy binding by other antibodies. One antibody was sensitive to changes in any of three separate parts of the molecule and some antibodies to distinct epitopes cross-blocked each other. Certain antibodies had the ability to increase HA binding by lymphocytes but this did not correlate absolutely with antibody specificity and was only partially attributable to CD44 cross-linking. Antibodies that consistently blocked HA recognition were all sensitive to amino acid changes within a short stretch of CD44. Such blocking antibodies interacted with CD44 more strongly than ligand in competition experiments. One large group of antibodies blocked ligand binding, but only with a particular cell line. This detailed analysis adds to our understanding of functional domains within CD44 and requirements for antibodies to influence recognition of one ligand.

scholarly journals Antigenic drift in influenza A viruses. I. Selection and characterization of antigenic variants of A/PR/8/34 [HON1] influenza virus with monoclonal antibodies

1978 ◽  
Vol 148 (2) ◽  
pp. 383-392 ◽  
Author(s):  
W Gerhard ◽  
RG Webster
Keyword(s):  
Influenza Virus ◽  
Monoclonal Antibodies ◽  
Amino Acid ◽  
Influenza A ◽  
Antigenic Drift ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Parent Virus ◽  
Influenza A Viruses

Antigenic variants of A/PR/8/34 [HON1] influenza virus were selected after a single passage of the parent virus in embryonated chicken eggs in the presence of monoclonal antibodies to this virus. The monoclonal antibodies were produced by a hybridoma and were specific for an antigenic determinant on the HA molecule of the parent virus. Seven antigenic variants were analyzed with 95 monoclonal anti-HA antibodies prepared in vitro in the splenic fragment culture system. Three subgroups of antigenic variants were distinguished. The antigenic changes were primarily recognized by monoclonal antibodies to the strain- specific determinants of the parental hemagglutinin (HA) molecule. Monoclonal antibodies to HA determinants shared (in an identical or cross-reactive form) by parental virus and more than three heterologous viruses of the HON1 and H1N1 subtypes were unable to recognize the antigenic change on the variants. Similarly, heterogeneous antibody preparations could not differentiate between parental and variant viruses. The results are compatible with the idea that the HA of PR8 has available a large repertoire of antigenic modifications that may result from single amino acid substitutions, and that antigenic changes can occur in the strain- specific determinants on the HA molecule in the absence of concomitant changes in the cross-reactive HA determinants. The findings suggest that antigenic drift, in order to be epidemiologically significant, probably requires a series of amino acid substitutions in, or close to, the antigenic area on the HA molecule.

scholarly journals A Single Point Mutation, Asn16→Lys, Dictates the Temperature-Sensitivity of the Reovirus tsG453 Mutant

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 289
Author(s):  
Kathleen K. M. Glover ◽  
Danica M. Sutherland ◽  
Terence S. Dermody ◽  
Kevin M. Coombs
Keyword(s):  
Amino Acid ◽  
Temperature Sensitivity ◽  
Reverse Genetics ◽  
Core Protein ◽  
Single Point ◽  
Single Amino Acid ◽  
Single Point Mutation ◽  
Temperature Sensitive ◽  
Amino Acid Substitutions ◽  
Gene Encoding

Studies of conditionally lethal mutants can help delineate the structure-function relationships of biomolecules. Temperature-sensitive (ts) mammalian reovirus (MRV) mutants were isolated and characterized many years ago. Two of the most well-defined MRV ts mutants are tsC447, which contains mutations in the S2 gene encoding viral core protein σ2, and tsG453, which contains mutations in the S4 gene encoding major outer-capsid protein σ3. Because many MRV ts mutants, including both tsC447 and tsG453, encode multiple amino acid substitutions, the specific amino acid substitutions responsible for the ts phenotype are unknown. We used reverse genetics to recover recombinant reoviruses containing the single amino acid polymorphisms present in ts mutants tsC447 and tsG453 and assessed the recombinant viruses for temperature-sensitivity by efficiency-of-plating assays. Of the three amino acid substitutions in the tsG453 S4 gene, Asn16-Lys was solely responsible for the tsG453ts phenotype. Additionally, the mutant tsC447 Ala188-Val mutation did not induce a temperature-sensitive phenotype. This study is the first to employ reverse genetics to identify the dominant amino acid substitutions responsible for the tsC447 and tsG453 mutations and relate these substitutions to respective phenotypes. Further studies of other MRV ts mutants are warranted to define the sequence polymorphisms responsible for temperature sensitivity.

scholarly journals Natural variants in SARS-CoV-2 Spike protein pinpoint structural and functional hotspots with implications for prophylaxis and therapeutic strategies

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Suman Pokhrel ◽  
Benjamin R. Kraemer ◽  
Scott Burkholz ◽  
Daria Mochly-Rosen
Keyword(s):  
Amino Acid ◽  
Severe Disease ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Severe Respiratory Distress ◽  
S Protein ◽  
Individual Sequences ◽  
Natural Variants ◽  
Novel Coronavirus ◽  
The Individual

AbstractIn December 2019, a novel coronavirus, termed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was identified as the cause of pneumonia with severe respiratory distress and outbreaks in Wuhan, China. The rapid and global spread of SARS-CoV-2 resulted in the coronavirus 2019 (COVID-19) pandemic. Earlier during the pandemic, there were limited genetic viral variations. As millions of people became infected, multiple single amino acid substitutions emerged. Many of these substitutions have no consequences. However, some of the new variants show a greater infection rate, more severe disease, and reduced sensitivity to current prophylaxes and treatments. Of particular importance in SARS-CoV-2 transmission are mutations that occur in the Spike (S) protein, the protein on the viral outer envelope that binds to the human angiotensin-converting enzyme receptor (hACE2). Here, we conducted a comprehensive analysis of 441,168 individual virus sequences isolated from humans throughout the world. From the individual sequences, we identified 3540 unique amino acid substitutions in the S protein. Analysis of these different variants in the S protein pinpointed important functional and structural sites in the protein. This information may guide the development of effective vaccines and therapeutics to help arrest the spread of the COVID-19 pandemic.

Single Amino Acid Substitutions Disrupt Tetramer Formation in the Dihydroneopterin Aldolase Enzyme ofPneumocystis carinii†

Biochemistry ◽  
1998 ◽  
Vol 37 (33) ◽  
pp. 11629-11636 ◽  
Author(s):  
M. Carmen Thomas ◽  
Stuart P. Ballantine ◽  
Susanne S. Bethell ◽  
Satty Bains ◽  
Paul Kellam ◽  
...  
Keyword(s):  
Amino Acid ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Tetramer Formation ◽  
Dihydroneopterin Aldolase

scholarly journals Pathogenic alleles in microtubule, secretory granule and extracellular matrix-related genes in familial keratoconus

2021 ◽  
Author(s):  
Vishal Shinde ◽  
Nara Sobreira ◽  
Elizabeth S Wohler ◽  
George Maiti ◽  
Nan Hu ◽  
...  
Keyword(s):  
Amino Acid ◽  
Cell Cultures ◽  
Cell Biology ◽  
Stromal Cell ◽  
Primary Cilia ◽  
Single Amino Acid ◽  
European Ancestry ◽  
Base Substitution ◽  
Amino Acid Substitutions ◽  
Functional Link

Abstract Keratoconus is a common corneal defect with a complex genetic basis. By whole exome sequencing of affected members from 11 multiplex families of European ancestry, we identified 23 rare, heterozygous, potentially pathogenic variants in 8 genes. These include nonsynonymous single amino acid substitutions in HSPG2, EML6 and CENPF in two families each, and in NBEAL2, LRP1B, PIK3CG and MRGPRD in three families each; ITGAX had nonsynonymous single amino acid substitutions in two families and an indel with a base substitution producing a nonsense allele in the third family. Only HSPG2, EML6 and CENPF have been associated with ocular phenotypes previously. With the exception of MRGPRD and ITGAX, we detected the transcript and encoded protein of the remaining genes in the cornea and corneal cell cultures. Cultured stromal cells showed cytoplasmic punctate staining of NBEAL2, staining of the fibrillar cytoskeletal network by EML6, while CENPF localized to the basal body of primary cilia. We inhibited the expression of HSPG2, EML6, NBEAL2 and CENPF in stromal cell cultures and assayed for the expression of COL1A1 as a readout of corneal matrix production. An upregulation in COL1A1 after siRNA inhibition indicated their functional link to stromal cell biology. For ITGAX, encoding a leukocyte integrin, we assayed its level in the sera of 3 affected families compared with 10 unrelated controls to detect an increase in all affecteds. Our study identified genes that regulate the cytoskeleton, protein trafficking and secretion, barrier tissue function and response to injury and inflammation, as being relevant to keratoconus.

scholarly journals Identification of separate domains in the adenovirus E1A gene for immortalization activity and the activation of virus early genes.

1986 ◽  
Vol 6 (10) ◽  
pp. 3470-3480 ◽  
Author(s):  
E Moran ◽  
B Zerler ◽  
T M Harrison ◽  
M B Mathews
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Point Mutations ◽  
Apparent Molecular Weight ◽  
Amino Acid Position ◽  
Cysteine Residue ◽  
Transformation Function ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
E1a Gene

The transformation and early adenovirus gene transactivation functions of the E1A region were analyzed with deletion and point mutations. Deletion of amino acids from position 86 through 120 had little effect on the lytic or transforming functions of the E1A products, while deletion of amino acids from position 121 through 150 significantly impaired both functions. The sensitivity of the transformation function to alterations in the region from amino acid position 121 to 150 was further indicated by the impairment of transforming activity resulting from single amino acid substitutions at positions 124 and 135. Interestingly, conversion of a cysteine residue at position 124 to glycine severely impaired the transformation function without affecting the early adenovirus gene activating functions. Single amino acid substitutions in a different region of the E1A gene had the converse effect. All the mutants produced polypeptides of sufficient stability to be detected by Western immunoblot analysis. The single amino acid substitutions at positions 124 and 135, although impairing the transformation functions, did not detectably alter the formation of the higher-apparent-molecular-weight forms of the E1A products.

scholarly journals Functional effects of single amino acid substitutions in the region of Phe113 to Asp138 in the plasminogen activator inhibitor 1 molecule

1998 ◽  
Vol 331 (2) ◽  
pp. 409-415 ◽  
Author(s):  
Guang-Chao SUI ◽  
Björn WIMAN
Keyword(s):  
Amino Acid ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Site Directed Mutagenesis ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Plasminogen Activator Inhibitor 1 ◽  
Low Activity ◽  
Activator Inhibitor ◽  
Pai 1

Thirteen amino acid substitutions have been introduced within the stretch Phe113 to Asp138 in the plasminogen activator inhibitor 1 (PAI-1) molecule by site-directed mutagenesis. The different proteins and wild-type (wt) PAI-1 have been overexpressed in Escherichia coliand purified by chromatography on heparin–Sepharose and on anhydrotrypsin–agarose. The PAI-1 variants have been characterized by their reactivity with tissue plasminogen activator (tPA), interactions with vitronectin or heparin, and stability. Most PAI-1 variants, except for Asp125 → Lys, Phe126 → Ser and Arg133 → Asp, displayed a high spontaneous inhibitory activity towards tPA, which did not change greatly on reactivation with 4 M guanidinium chloride, followed by dialysis at pH 5.5. The variants Asp125 → Lys and Arg133 → Asp became much more active after reactivation and they were also more rapidly transformed to inactive forms (t½ 22–31 min) at physiological pH and temperature than the other variants. However, in the presence of vitronectin they were both almost equally stable (t½ 2.3 h) as wtPAI-1 (t½ 3.0 h). The mutant Glu130 → Lys showed an increased stability, both in the absence and in the presence of vitronectin compared with wtPAI-1. Nevertheless a similar affinity between all the active PAI-1 variants and vitronectin was observed. Further, all mutants, including the three mutants with low activity, were to a large extent adsorbed on anhydrotrypsin–agarose and were eluted in a similar fashion. In accordance with these data, the three variants with a low activity were all to a large extent cleaved as a result of their reaction with tPA, suggesting that they occurred predominantly in the substrate conformation. Our results do not support the presence of a binding site for vitronectin in this part of the molecule, but rather that it might be involved in controlling the active PAI-1 to substrate transition. Partly, this region of the PAI-1 molecule (Arg115 to Arg118) seems also to be involved in the binding of heparin to PAI-1.

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