Abstract:
:Rational engineering methods can be applied with reasonable success to optimize physicochemical characteristics of proteins, in particular, antibodies. Here, we describe a combined CDR3 walking randomization and rational design-based approach to enhance the affinity of the human anti-gastrin TA4 scFv. The application of this methodology to TA4 scFv, displaying only a weak overall affinity for gastrin17 (K(D) = 6 microM), resulted in a set of nine affinity-matured scFv variants with near-nanomolar affinity (K(D) = 13.2 nM for scFv TA4.112). First, CDR-H3 and CDR-L3 randomization resulted in three scFvs with an overall affinity improvement of 15- to 35-fold over the parental. Then, the modeling of two scFv constructs selected from the previous step (TA4.11 and TA4.13) was followed by a combination of manual and molecular dynamics-based docking of gastrin17 into the respective binding sites, analysis of apparent packing defects, and selection of residues for mutagenesis through phage display. Nine scFv mutants were obtained from the second maturation step. A final 454-fold improvement in affinity compared with TA4 was obtained. These scFvs showed an enhanced potency to inhibit gastrin-induced proliferation in Colo 320 WT and BxPc3 tumoral cells. In conclusion, we propose a structure-based rational method to accelerate the development of affinity-matured antibody constructs with enhanced potential for therapeutic use.
journal_name
Proc Natl Acad Sci U S Aauthors
Barderas R,Desmet J,Timmerman P,Meloen R,Casal JIdoi
10.1073/pnas.0801221105subject
Has Abstractpub_date
2008-07-01 00:00:00pages
9029-34issue
26eissn
0027-8424issn
1091-6490pii
0801221105journal_volume
105pub_type
杂志文章abstract::We exploit the unusual genome organization of the ciliate cell to analyze the control of specific gene amplification during a nuclear differentiation process. Ciliates contain two types of nuclei within one cell, the macronucleus and the micronucleus; and after sexual reproduction a new macronucleus is formed from a m...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1009284107
更新日期:2010-12-21 00:00:00
abstract::By using harmonic radar, we report the complete flight paths of displaced bees. Test bees forage at a feeder or are recruited by a waggle dance indicating the feeder. The flights are recorded after the bees are captured when leaving the hive or the feeder and are released at an unexpected release site. A sequence of b...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.0408550102
更新日期:2005-02-22 00:00:00
abstract::The blind Mexican cave tetra, Astyanax mexicanus, is a unique model system for the study of parallelism and the evolution of cave-adapted traits. Understanding the genetic basis for these traits has recently become feasible thanks to production of a genome-wide linkage map and quantitative trait association analyses. ...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.0806238105
更新日期:2008-12-23 00:00:00
abstract::Vesicles were constituted with glycophorin, the Sendai virus receptor of human erythrocytes, and loaded with calcein, a polar derivative of fluorescein, at self-quenching concentrations. On exposure to Sendai virus and mild hypo-osmotic stress, vesicles of the appropriate composition released a significant portion of ...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.80.6.1608
更新日期:1983-03-01 00:00:00
abstract::Peptide nucleic acid (PNA) is a DNA mimic that has shown considerable promise as a lead compound for developing gene therapeutic drugs. We report that PNAs targeted to functional and accessible sites in ribosomal RNA can inhibit translation in an Escherichia coli cell-free transcription/translation system, with 50% re...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.95.5.2073
更新日期:1998-03-03 00:00:00
abstract::IL-33, a cytokine of the IL-1 family, is closely associated with type II T cell responses. Here, we report an unexpected proinflammatory role of IL-33 in inflammatory arthritis. IL-33 was expressed in synovial fibroblasts from patients with rheumatoid arthritis (RA). Expression was markedly elevated in vitro by inflam...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.0801898105
更新日期:2008-08-05 00:00:00
abstract::The DNA of functional cis-regulatory modules displays extensive sequence conservation in comparisons of genomes from modestly distant species. Patches of sequence that are several hundred base pairs in length within these modules are often seen to be 80-95% identical, although the flanking sequence cannot even be alig...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.0505291102
更新日期:2005-08-16 00:00:00
abstract::Antischizophrenic neuroleptic drugs of the diphenylbutylpiperidine class, which includes pimozide, fluspirilene, penfluridol, and clopimozide, inhibit [3H]nitrendipine binding with IC50 values of 13-30 nM. This inhibition involves receptors for the verapamil/prenylamine class of calcium channel antagonists. These diph...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.80.16.5122
更新日期:1983-08-01 00:00:00
abstract::Two-dimensional gel electrophoresis of immunoprecipitated human HLA-DR antigens from cells expressing the HLA-DR4 haplotype shows distinct clustering of beta-chain patterns. Six unique electrophoretic variants were observed among 17 HLA-DR4 homozygous cell lines (HCL) analyzed. These patterns correlate precisely with ...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.80.22.6962
更新日期:1983-11-01 00:00:00
abstract::The usual bias in earthquake catalogs against shocks of small magnitudes can be removed by testing the randomness of the magnitudes of successive shocks. The southern California catalog, 1933-1967, is found to be unbiased in the sense of the test at magnitude 4 or above; the cutoff is improved to M = 3 for the subcata...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.63.4.1051
更新日期:1969-08-01 00:00:00
abstract::During the early evolution of life, 8-oxo-7,8-dihydro-2'-deoxyguanosine (O) may have functioned as a proto-flavin capable of repairing cyclobutane pyrimidine dimers in DNA or RNA by photoinduced electron transfer using longer wavelength UVB radiation. To investigate the ability of O to act as an excited-state electron...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1404411111
更新日期:2014-08-12 00:00:00
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journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.0500619102
更新日期:2005-03-15 00:00:00
abstract::The strikingly high incidence of obstructed labor due to the disproportion of fetal size and the mother's pelvic dimensions has puzzled evolutionary scientists for decades. Here we propose that these high rates are a direct consequence of the distinct characteristics of human obstetric selection. Neonatal size relativ...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1612410113
更新日期:2016-12-20 00:00:00
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journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.94.10.5006
更新日期:1997-05-13 00:00:00
abstract::In eukaryotes, ubiquitination is an important posttranslational process achieved through a cascade of ubiquitin-activating (E1), conjugating (E2), and ligase (E3) enzymes. Many pathogenic bacteria deliver virulence factors into the host cell that function as E3 ligases. How these bacterial "Trojan horses" integrate in...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1115025109
更新日期:2012-02-07 00:00:00
abstract::The primary structure of the beta chain of human glycoprotein Ib (GPIb), the platelet receptor for von Willebrand factor, has been established by a combination of cDNA cloning and amino acid sequence analysis. A lambda phage cDNA expression library prepared from human erythroleukemia cells (HEL cells) was screened wit...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.85.7.2135
更新日期:1988-04-01 00:00:00
abstract::Tumor-derived adhesion factor (TAF) was previously identified as a cell adhesion molecule secreted by human bladder carcinoma cell line EJ-1. To elucidate the physiological function of TAF, we examined its distribution in human normal and tumor tissues. Immunochemical staining with an anti-TAF monoclonal antibody show...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.93.16.8384
更新日期:1996-08-06 00:00:00
abstract::Although the first observations of the inhibitory effect of methylation on gene activity were made almost a decade ago, the mechanism by which methyl groups affect transcription is still obscure. Here we use engineered promoters methylated in vitro in transient transfections to study the mechanism by which methylation...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.89.21.10119
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abstract::Epstein-Barr virus (EBV)-genome-negative human lymphoma lines, Ramos and BJAB, can be converted by EBV in vitro into EBV-genome-positive virus nonproducer lines. These cell lines have been used to identify surface antigens unique to EBV, with the expectation that such determinants might be related to the antigenic tar...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
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journal_title:Proceedings of the National Academy of Sciences of the United States of America
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更新日期:2001-03-27 00:00:00
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journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.79.15.4645
更新日期:1982-08-01 00:00:00
abstract::Bone remodeling requires osteoclasts to generate and maintain an acidified resorption compartment between the apical membrane and the bone surface to solubilize hydroxyapatite crystals within the bone matrix. This acidification process requires (i) apical proton secretion by a vacuolar H(+)-ATPase, (ii) actin cytoskel...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1206392110
更新日期:2013-02-05 00:00:00
abstract::A galanin receptor protein was solubilized with 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) from pig brain membranes and then purified by single-step affinity chromatography. The product exhibits saturable and specific binding for galanin with a binding activity of 17 nmol/mg of protein and a dis...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.90.9.3845
更新日期:1993-05-01 00:00:00
abstract::Forkhead box g1 (Foxg1) is a nuclear-cytosolic transcription factor essential for the forebrain development and involved in neurodevelopmental and cancer pathologies. Despite the importance of this protein, little is known about the modalities by which it exerts such a large number of cellular functions. Here we show ...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1515190112
更新日期:2015-11-10 00:00:00
abstract::The enzymes responsible for the oxidation of n-octane to octanoic acid or beyond in Pseudomonas oleovorans are octane inducible and are coded by genes borne on a transmissible extrachromosomal element. The octane to octanoate enzymes induced by octane are repressed by octanol. The chromosome also carries genes coding ...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.70.4.1137
更新日期:1973-04-01 00:00:00
abstract::The study of proteolysis lies at the heart of our understanding of biocatalysis, enzyme evolution, and drug development. To understand the degree of natural variation in protease active sites, we systematically evaluated simple active site features from all serine, cysteine and threonine proteases of independent linea...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1221050110
更新日期:2013-02-19 00:00:00
abstract::In the nitrogen-fixing symbiosis between legume hosts and rhizobia, the bacteria are engulfed by a plant cell membrane to become intracellular organelles. In the model legume Medicago truncatula, internalization and differentiation of Sinorhizobium (also known as Ensifer) meliloti is a prerequisite for nitrogen fixati...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1500123112
更新日期:2015-12-08 00:00:00
abstract::While cells within tissues generate and sense 3D states of strain, the current understanding of the mechanics of fibrous extracellular matrices (ECMs) stems mainly from uniaxial, biaxial, and shear tests. Here, we demonstrate that the multiaxial deformations of fiber networks in 3D cannot be inferred solely based on t...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1815659116
更新日期:2019-04-02 00:00:00
abstract::Human cells express thousands of different surface proteins that can be used for cell classification, or to distinguish healthy and disease conditions. A method capable of profiling a substantial fraction of the surface proteome simultaneously and inexpensively would enable more accurate and complete classification of...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.1721899115
更新日期:2018-03-13 00:00:00
abstract::Electrophilic neurite outgrowth-promoting prostaglandin (NEPP) compounds protect neurons from oxidative insults. At least part of the neuroprotective action of NEPPs lies in induction of hemeoxygenase-1 (HO-1), which, along with other phase II enzymes, serve as a defense system against oxidative stress. Here, we found...
journal_title:Proceedings of the National Academy of Sciences of the United States of America
pub_type: 杂志文章
doi:10.1073/pnas.0505723102
更新日期:2006-01-17 00:00:00