scholarly journals Crystal Structures and Functional Characterization of Wild-Type CYP101D1 and Its Active Site Mutants

Biochemistry ◽  
2013 ◽  
Vol 52 (49) ◽  
pp. 8898-8906 ◽  
Author(s):  
Dipanwita Batabyal ◽  
Thomas L. Poulos
Keyword(s):  
Crystal Structures ◽  
Active Site ◽  
Functional Characterization ◽  
Wild Type ◽  
Active Site Mutants

Functional characterization of the H+/K+ ATPase in wild type and gastrin knock-out mice

2007 ◽  
Vol 45 (05) ◽  
Author(s):  
A Schnur ◽  
P Hegyi ◽  
V Venglovecz ◽  
Z Rakonczay ◽  
I Ignáth ◽  
...  
Keyword(s):  
Functional Characterization ◽  
Wild Type ◽  
Knock Out ◽  
Knock Out Mice

scholarly journals Expression and characterization of active site mutants of hevamine, a chitinase from the rubber tree Hevea brasiliensis

2002 ◽  
Vol 269 (3) ◽  
pp. 893-901 ◽  
Author(s):  
Evert Bokma ◽  
Henriëtte J. Rozeboom ◽  
Mark Sibbald ◽  
Bauke W. Dijkstra ◽  
Jaap J. Beintema
Keyword(s):  
Active Site ◽  
Hevea Brasiliensis ◽  
Rubber Tree ◽  
Active Site Mutants

scholarly journals Crystal Structures of E. coli Native MenH and Two Active Site Mutants

PLoS ONE ◽  
2013 ◽  
Vol 8 (4) ◽  
pp. e61325 ◽  
Author(s):  
Jodie M. Johnston ◽  
Ming Jiang ◽  
Zhihong Guo ◽  
Edward N. Baker
Keyword(s):  
Crystal Structures ◽  
Active Site ◽  
E Coli ◽  
Active Site Mutants

scholarly journals Functional characterization of the rod visual pigment of the echidna (Tachyglossus aculeatus), a basal mammal

2012 ◽  
Vol 29 (4-5) ◽  
pp. 211-217 ◽  
Author(s):  
CONSTANZE BICKELMANN ◽  
JAMES M. MORROW ◽  
JOHANNES MÜLLER ◽  
BELINDA S.W. CHANG
Keyword(s):  
Half Life ◽  
Functional Characterization ◽  
Egg Laying ◽  
Sensory Transduction ◽  
Wild Type ◽  
Functional Variation ◽  
Bovine Rhodopsin ◽  
Tachyglossus Aculeatus

AbstractMonotremes are the most basal egg-laying mammals comprised of two extant genera, which are largely nocturnal. Visual pigments, the first step in the sensory transduction cascade in photoreceptors of the eye, have been examined in a variety of vertebrates, but little work has been done to study the rhodopsin of monotremes. We isolated the rhodopsin gene of the nocturnal short-beaked echidna (Tachyglossus aculeatus) and expressed and functionally characterized the protein in vitro. Three mutants were also expressed and characterized: N83D, an important site for spectral tuning and metarhodopsin kinetics, and two sites with amino acids unique to the echidna (T158A and F169A). The λmax of echidna rhodopsin (497.9 ± 1.1 nm) did not vary significantly in either T158A (498.0 ± 1.3 nm) or F169A (499.4 ± 0.1 nm) but was redshifted in N83D (503.8 ± 1.5 nm). Unlike other mammalian rhodopsins, echidna rhodopsin did react when exposed to hydroxylamine, although not as fast as cone opsins. The retinal release rate of light-activated echidna rhodopsin, as measured by fluorescence spectroscopy, had a half-life of 9.5 ± 2.6 min−1, which is significantly shorter than that of bovine rhodopsin. The half-life of the N83D mutant was 5.1 ± 0.1 min−1, even shorter than wild type. Our results show that with respect to hydroxylamine sensitivity and retinal release, the wild-type echidna rhodopsin displays major differences to all previously characterized mammalian rhodopsins and appears more similar to other nonmammalian vertebrate rhodopsins such as chicken and anole. However, our N83D mutagenesis results suggest that this site may mediate adaptation in the echidna to dim light environments, possibly via increased stability of light-activated intermediates. This study is the first characterization of a rhodopsin from a most basal mammal and indicates that there might be more functional variation in mammalian rhodopsins than previously assumed.

scholarly journals Functional characterization of wild-type and mutant human sialin

2004 ◽  
Vol 23 (23) ◽  
pp. 4560-4570 ◽  
Author(s):  
Pierre Morin ◽  
Corinne Sagné ◽  
Bruno Gasnier
Keyword(s):  
Functional Characterization ◽  
Wild Type

scholarly journals 2066 Functional characterization of mutant BRCA1

2018 ◽  
Vol 2 (S1) ◽  
pp. 13-13
Author(s):  
John Barrows ◽  
David Long
Keyword(s):  
Functional Characterization ◽  
Coiled Coil ◽  
Wild Type ◽  
Coiled Coil Region ◽  
Egg Extracts ◽  
Study Population ◽  
Xenopus Egg ◽  
Xenopus Egg Extracts

OBJECTIVES/SPECIFIC AIMS: The objective of this work is to determine the mechanistic consequences of BRCA1 mutants in inter-strand crosslink (ICL) repair. METHODS/STUDY POPULATION: Our lab uses Xenopus egg extracts to study ICL repair. These extracts can be depleted of endogenous BRCA1 by immunoprecipitation. The goal of this work is to rescue endogenous depletion with in vitro translated, wild type BRCA1. Once achieved, we can supplement the depleted extract with BRCA1 mutants to access their function in ICL repair. RESULTS/ANTICIPATED RESULTS: We hypothesize that the BRCT and RING domain mutations will abrogate ICL repair, while mutations in the coiled coil region will not affect repair. DISCUSSION/SIGNIFICANCE OF IMPACT: These findings will have an immense impact on the understanding of BRCA1 domains. Importantly these results will spur personalized therapy of BRCA1 mutants by showing which domains are sensitive to cross-linking agents.

scholarly journals Structural and Functional Characterization of the Human Thymidylate Synthase (hTS) Interface Variant R175C, New Perspectives for the Development of hTS Inhibitors

Molecules ◽  
2019 ◽  
Vol 24 (7) ◽  
pp. 1362
Author(s):  
Cecilia Pozzi ◽  
Stefania Ferrari ◽  
Rosaria Luciani ◽  
Maria Costi ◽  
Stefano Mangani
Keyword(s):  
Thymidylate Synthase ◽  
Active Site ◽  
Functional Characterization ◽  
Binding Mode ◽  
Anticancer Chemotherapy ◽  
Innovative Strategies ◽  
X Ray Crystallography ◽  
Site Targeting ◽  
New Strategies

Human thymidylate synthase (hTS) is pivotal for cell survival and proliferation, indeed it provides the only synthetic source of dTMP, required for DNA biosynthesis. hTS represents a validated target for anticancer chemotherapy. However, active site-targeting drugs towards hTS have limitations connected to the onset of resistance. Thus, new strategies have to be applied to effectively target hTS without inducing resistance in cancer cells. Here, we report the generation and the functional and structural characterization of a new hTS interface variant in which Arg175 is replaced by a cysteine. Arg175 is located at the interface of the hTS obligate homodimer and protrudes inside the active site of the partner subunit, in which it provides a fundamental contribution for substrate binding. Indeed, the R175C variant results catalytically inactive. The introduction of a cysteine at the dimer interface is functional for development of new hTS inhibitors through innovative strategies, such as the tethering approach. Structural analysis, performed through X-ray crystallography, has revealed that a cofactor derivative is entrapped inside the catalytic cavity of the hTS R175C variant. The peculiar binding mode of the cofactor analogue suggests new clues exploitable for the design of new hTS inhibitors.

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