bluuebunny/crossref_metadata_2025_split huggingface dataset:
CYP94A5, a new cytochrome P450 from Nicotiana tabacum is able to catalyze the oxidation of fatty acids to the ω‐alcohol and to the corresponding diacid
Authors: Le Bouquin Renaud , Skrabs Manuela , Kahn Rachel , Benveniste Irène , Salaün Jean‐Pierre , Schreiber Lukas , Durst Francis , Pinot Franck 
Published: March 2003

A full length cDNA encoding a new cytochrome P450‐dependent fatty acid hydroxylase (CYP94A5) was isolated from a tobacco cDNA library. CYP94A5 was expressed in S. cerevisiae strain WAT11 containing a P450 reductase from Arabidopsis thaliana necessary for catalytic activity of cytochrome P450 enzymes. When incubated for 10 min in presence of NADPH with microsomes of recombinant yeast, 9,10‐epoxystearic acid was converted into one major metabolite identified by GC/MS as 18‐hydroxy‐9,10‐epoxystearic acid. The kinetic parameters of the reaction were Km,app = 0.9 ± 0.2 µm and Vmax,app = 27 ± 1 nmol·min−1·nmol−1 P450. Increasing the incubation time to 1 h led to the formation of a compound identified by GC/MS as 9,10‐epoxy‐octadecan‐1,18‐dioic acid. The diacid was also produced in microsomal incubations of 18‐hydroxy‐9,10‐epoxystearic acid. Metabolites were not produced in incubations with microsomes of yeast transformed with a control plasmid lacking CYP94A5 and their production was inhibited by antibodies raised against the P450 reductase, demonstrating the involvement of CYP94A5 in the reactions. The present study describes a cytochrome P450 able to catalyze the complete set of reactions oxidizing a terminal methyl group to the corresponding carboxyl. This new fatty acid hydroxylase is enantioselective: after incubation of a synthetic racemic mixture of 9,10‐epoxystearic acid, the chirality of the residual epoxide was 40/60 in favor of 9R,10S enantiomer. CYP94A5 also catalyzed the ω‐hydroxylation of saturated and unsaturated fatty acids with aliphatic chain ranging from C12 to C18.
Detection of a tryptophan radical in the reaction of ascorbate peroxidase with hydrogen peroxide
Authors: Hiner Alexander N. P. , Martínez Jesús I. , Arnao Marino B. , Acosta Manuel , Turner Daniel D. , Lloyd Raven Emma , Rodríguez‐López José Neptuno 
Published: March 2003

The reactivity of recombinant pea cytosolic ascorbate peroxidase (rAPX) towards H2O2, the nature of the intermediates and the products of the reaction have been examined using UV/visible and EPR spectroscopies together with HPLC. Compound I of rAPX, generated by reaction of rAPX with 1 molar equivalent of H2O2, contains a porphyrin π‐cation radical. This species is unstable and, in the absence of reducing substrate, decays within 60 s to a second species, compound I*, that has a UV/visible spectrum [λmax (nm) = 414, 527, 558 and 350 (sh)] similar, but not identical, to those of both horseradish peroxidase compound II and cytochrome c peroxidase compound I. Small but systematic differences were observed in the UV/visible spectra of compound I* and authentic rAPX compound II, generated by reaction of rAPX with 1 molar equivalent H2O2 in the presence of 1 molar equivalent of ascorbate [λmax (nm) = 416, 527, 554, 350 (sh) and 628 (sh)]. Compound I* decays to give a ‘ferric‐like’ species (λmax = 406 nm) that is not spectroscopically identical to ferric rAPX (λmax = 403 nm) with a first order rate constant, kdecay′ = (2.7 ± 0.3) × 10−4 s−1. Authentic samples of compound II evolve to ferric rAPX [kdecay = (1.1 ± 0.2) × 10−3 s−1]. Low temperature (10 K) EPR spectra are consistent with the formation of a protein‐based radical, with g values for compound I* (g∥ = 2.038, g⊥ = 2.008) close to those previously reported for the Trp191 radical in cytochrome c peroxidase (g∥ = 2.037, g⊥ = 2.005). The EPR spectrum of rAPX compound II was essentially silent in the g = 2 region. Tryptic digestion of the ‘ferric‐like’ rAPX followed by RP‐HPLC revealed a fragment with a new absorption peak near 330 nm, consistent with the formation of a hydroxylated tryptophan residue. The results show, for the first time, that rAPX can, under certain conditions, form a protein‐based radical analogous to that found in cytochrome c peroxidase. The implications of these data are discussed in the wider context of both APX catalysis and radical formation and stability in haem peroxidases.
Regulation of cellular senescence by p53
Authors: Itahana Koji , Dimri Goberdhan , Campisi Judith 
Published: March 2003

Many normal cells respond to potentially oncogenic stimuli by undergoing cellular senescence, a state of irreversibly arrested proliferation and altered differentiated function. Cellular senescence very likely evolved to suppress tumorigenesis. In support of this idea, it is regulated by several tumor suppressor genes. At the heart of this regulation is p53. p53 is essential for the senescence response to short telomeres, DNA damage, oncogenes and supraphysiological mitogenic signals, and overexpression of certain tumor suppressor genes. Despite the well‐documented central role for p53 in the senescence response, many questions remain regarding how p53 senses senescence‐inducing stimuli and how it elicits the senescent phenotype.
Methionine oxidation and its effect on the stability of a reconstituted subunit of the light‐harvesting complex from Rhodospirillum rubrum
Authors: Wang Zheng‐Yu , Shimonaga Masahiro , Muraoka Yoshiyuki , Kobayashi Masayuki , Nozawa Tsunenori 
Published: March 2003

An additional component in the purified core light‐harvesting complex (LH1) from wild‐type purple photosynthetic bacterium Rhodospirillum rubrum has been identified as an oxidized species of α‐polypeptide by MALDI‐TOF mass spectrometry. This component appears as a slightly earlier‐eluting peak in the RP‐HPLC chromatogram compared with the authentic α‐polypeptide. The oxidation site has been determined to be the N‐terminal methionine residue by high‐resolution NMR spectroscopy, where the methionine is oxidized to methionine sulfoxide in a diastereoisomeric form. Interconversion between the oxidized and authentic α‐polypeptides has been confirmed by selective oxidation and reduction. The oxidative modification of methionine is shown to have discernible effects on the ability to form B820 subunit with β‐polypeptide and bacteriochlorophyll a, and on the stability of the reconstituted B820 subunit. Both the ability and the stability for the samples using the oxidized α‐polypeptide are moderately reduced, indicating that the oxidation‐induced conformational change in the N‐terminal domain of α‐polypeptide may affect the pigment‐binding environment through a long‐range interaction. The MALDI‐TOF mass results also reveal that the N‐terminus of α‐polypeptide is formylated and no phosphorylation has occurred in this polypeptide.
Oxidative folding of murine prion mPrP(23–231)
Authors: Lu Bao‐Yuan , Beck Pamela J. , Chang Jui‐Yoa 
Published: March 2003

A systematic study of the oxidative folding of murine prion protein mPrP(23–231) is reported here. Folding of mPrP(23–231) involves formation of a single disulfide bond, Cys179–Cys214. Despite this simplicity, reduced mPrP(23–231) exhibits numerous unusual folding properties. In the absence of denaturant, folding of mPrP(23–231) is extremely sluggish, regardless of pH. The optimal pH for mPrP(23–231) folding was found to be 4–5. At pH 8.0, a condition that typically favors disulfide formation, folding of mPrP(23–231) hardly occurs, and it not facilitated by inclusion of redox agent. In the presence of denaturant (4 m urea or 2 m guanidine hydrochloride) and basic pH (8.0), reduced mPrP(23–231) refolds to the native structure quantitatively. The efficiency of folding can be further promoted by the presence of oxidized glutathione. At pH 4.0 and in the presence of 4 m urea, reduced mPrP(23–231) converts to three distinctive conformational isomers, unable to form the native structure. These unusual properties lead us to the following conclusions. The reduced mPrP(23–231) adopts a highly rigid structure with the two cysteines buried or situated apart. The presence of denaturant or low pH disrupts this rigid structure and lowers the energy barrier, which permits oxidation and refolding of the reduced mPrP(23–231). Under selected conditions, reduced mPrP(23–231) is capable of taking on multiple forms of stable conformational isomer that are segregated by energy barriers.
Binding interactions between barley thaumatin‐like proteins and (1,3)‐β‐D‐glucans
Authors: Osmond Ronald I. W. , Hrmova Maria , Fontaine Fabien , Imberty Anne , Fincher Geoffrey B. 
Published: March 2003

The specificity and kinetics of the interaction between the pathogenesis‐related group of thaumatin‐like proteins (PR5) in higher plants and (1,3)‐β‐d‐glucans have been investigated. Two thaumatin‐like proteins with 60% amino‐acid sequence identity were purified from extracts of germinated barley grain, and were designated HvPR5b and HvPR5c. Purified HvPR5c interacted with insoluble (1,3)‐β‐d‐glucans, but not with cellulose, pustulan, xylan, chitin or a yeast mannoprotein. Tight binding was observed with unbranched and unsubstituted (1,3)‐β‐d‐glucans, and weaker binding was seen if (1,6)‐β‐linked branch points or β‐glucosyl substituents were present in the substrate. The HvPR5b protein interacted weakly with insoluble (1,3)‐β‐d‐glucans and did not bind to any of the other polysaccharides tested. This indicated that only specific barley PR5 isoforms interact tightly with (1,3)‐β‐d‐glucans. The complete primary structures of HvPR5b and HvPR5c were determined and used to construct molecular models of HvPR5b and HvPR5c, based on known three‐dimensional structures of related thaumatin‐like proteins. The models were examined for features that may be associated with (1,3)‐β‐d‐glucan binding, and a potential (1,3)‐β‐d‐glucan‐binding region was located on the surface of HvPR5c. No obvious structural features that would prevent binding of (1,3)‐β‐d‐glucan to HvPR5b were identified, but several of the amino acids in HvPR5c that are likely to interact with (1,3)‐β‐d‐glucans are not present in HvPR5b.
Structural characterization of the C2 domain of novel protein kinase Cε
Authors: García‐García Josefa , Gómez‐Fernández Juan C. , Corbalán‐García Senena 
Published: March 2003

Infrared spectroscopy (IR) and differential scanning calorimetry (DSC) were used to study the biophysical properties of the PKCε‐C2 domain, a C2 domain that possess special characteristics as it binds to acidic phospholipids in a Ca2+‐independent manner and no structural information about it is available to date. When the secondary structure was determined by IR spectroscopy in H2O and D2O buffers, β sheet was seen to be the major structural component. Spectroscopic studies of the thermal denaturation in D2O showed a broadening in the amide I′ band starting at 45 °C. Curve fitting analysis of the spectra demonstrated that two components appear upon thermal denaturation, one at 1623 cm−1 which was assigned to aggregation and a second one at 1645 cm−1, which was assigned to unordered or open loop structures. A lipid binding assay has demonstrated that PKCε‐C2 domain has preferencial affinity for PIP2 although it exhibits maximal binding activity for phosphatidic acid when 100 mol% of this negatively charged phospholipid was used. Thus, phosphatidic acid containing vesicles were used to characterize the effect of lipid binding on the secondary structure and thermal stability. These experiments showed that the secondary structure did not change upon lipid binding and the thermal stability was very high with no significant changes occurring in the secondary structure after heating. DSC experiments demonstrated that when the C2‐protein was scanned alone, it showed a Tm of 49 °C and a calorimetric denaturation enthalpy of 144.318 kJ·mol−1. However, when phoshatidic acid vesicles were included in the mixture, the transition disappeared and further IR experiments demonstrated that the protein structure was not modified under these conditions.
Binding of Thermomyces (Humicola) lanuginosa lipase to the mixed micelles of cis‐parinaric acid/NaTDC
Authors: Yapoudjian Stéphane , Ivanova Margarita G. , Brzozowski A. Marek , Patkar Shamkant A. , Vind Jesper , Svendsen Allan , Verger Robert 
Published: March 2003

The binding of Thermomyces lanuginosa lipase and its mutants [TLL(S146A), TLL(W89L), TLL(W117F, W221H, W260H)] to the mixed micelles of cis‐parinaric acid/sodium taurodeoxycholate at pH 5.0 led to the quenching of the intrinsic tryptophan fluorescence emission (300–380 nm) and to a simultaneous increase in the cis‐parinaric acid fluorescence emission (380–500 nm). These findings were used to characterize the Thermomyces lanuginosa lipase/cis‐parinaric acid interactions occurring in the presence of sodium taurodeoxycholate.The fluorescence resonance energy transfer and Stern–Volmer quenching constant values obtained were correlated with the accessibility of the tryptophan residues to the cis‐parinaric acid and with the lid opening ability of Thermomyces lanuginosa lipase (and its mutants). TLL(S146A) was found to have the highest fluorescence resonance energy transfer.In addition, a TLL(S146A)/oleic acid complex was crystallised and its three‐dimensional structure was solved. Surprisingly, two possible binding modes (sn‐1 and antisn1) were found to exist between oleic acid and the catalytic cleft of the open conformation of TLL(S146A). Both binding modes involved an interaction with tryptophan 89 of the lipase lid, in agreement with fluorescence resonance energy transfer experiments.As a consequence, we concluded that TLL(S146A) mutant is not an appropriate substitute for the wild‐type Thermomyces lanuginosa lipase for mimicking the interaction between the wild‐type enzyme and lipids.
Microgeographic Variation and Sharing of the Gargle Vocalization and Its Component Syllables in Black‐Capped Chickadee (Aves, Paridae, Poecile atricapillus) Populations
Authors: Baker Myron C. , Howard Trina M. , Sweet Paul W. 
Published: March 2003

AbstractThroughout the year during agonistic encounters, black‐capped chickadees (Poecile atricapillus) emit a vocal signal known as the gargle call. Each bird has a repertoire of structurally differing gargle calls; some are shared with others in the local area. As a basis for understanding the cultural evolution of this social signal, we initiated a study of gargle call repertoires of birds living in a narrow belt of continuous riparian habitat occupied throughout by a resident population of chickadees. During two consecutive winter seasons, we sampled repertoires at three locations over a distance of 8.4 km to quantify micro‐geographical variation. Analyses of vocal sharing and population differentiation were carried out on whole gargle calls and on the individual acoustic units (syllables) from which the whole calls are constructed. We analysed 28 380 calls of 46 subjects in the two seasons of study. Birds averaged 7.6 different calls in their gargle repertoires. Calls were composed of about 10 syllables on average. Fifty‐six different syllables were used to construct the calls of all birds. Each study site had some gargle calls unique to the local birds and some that were shared with one or both of the other two sites. There was significantly greater sharing of both calls and syllables among birds within sample sites than between sample sites. The frequencies of the different kinds of gargles and syllables were significantly correlated across the 2 yr of the study, but the correlation was stronger (r2 = 0.93) for syllables than for whole gargle calls (r2 = 0.61).
Changes In Rhesus Macaque ‘Coo’ Vocalizations during Early Development
Authors: Hammerschmidt Kurt , Newman John D. , Champoux Maribeth , Suomi Stephen J. 
Published: March 2003

AbstractIn order to test whether ‘coo’ calls of young rhesus macaques, Macaca mulatta, undergo some modifications during early development, and to explore which factors may influence these changes, we studied the ontogeny of their contact call, the ‘coo’ call. Vocalizations were recorded during brief periods of social separation. Infants were either raised with their mothers and other conspecifics, or separated from their mothers at birth and housed in a nursery with other infants. We recorded calls uttered in the separation context from 20 infants. We digitized the first 50 calls of a given series and subjected them to a Fourier transform. From each frequency–time spectrum, we extracted 65 acoustic parameters using a software program (LMA 5.9). We then used a cluster analysis to separate the ‘coo’ calls from other call types. With increasing age, the ‘coos’ dropped in pitch and became more even. The course of amplitude became more constant and the call duration increased slightly. Nevertheless, we found a high intra‐individual variation throughout the 5 mo. Neither rearing condition nor sex had any apparent influence on age‐related changes in ‘coo’ structure. With one exception, all parameters that correlated with age could be explained by variation in weight. Therefore, we conclude that growth is the main factor accounting for the observed changes.
Consortship and Mating Success in Chacma Baboons (Papio cynocephalus ursinus)
Authors: Weingrill T. , Lycett J. E. , Henzi S. P. 
Published: March 2003

AbstractChacma baboons (Papio cynocephalus ursinus) show a lower consortship take‐over rate and longer consortship duration than the other savannah baboons (Bulger 1993). It has been argued that researchers have focused on atypically small troops with few adult males, resulting in low competition for access to oestrous females. Consortship data from two mountain baboon troops containing seven and four males, respectively, were analysed to determine whether the troop with the greater number of males showed a weaker correlation between mating success and rank due to an expected higher consortship take‐over rate. No consort take‐overs were observed in either study troop and mating success in both troops was correlated strongly with male rank. The distribution of days spent in consortship amongst the males could be explained by the priority‐of‐access‐model. The degree of cycle overlap determined the number of males observed consorting oestrous females, whereas the number of males did not influence the relationship between rank and consorting activity.
Insemination Capacity and Dispersal in Relation to Sex Allocation Decisions in Goniozus legneri (Hymenoptera: Bethylidae): Why Are There More Males in Larger Broods?
Authors: Hardy Ian C. W. , Stokkebo Stinne , Bønløkke‐Pedersen Jesper , Sejr Mikael K. 
Published: March 2003

AbstractModels considering sex ratio optima under single foundress strict local mate competition predict that female bias will be reduced by stochasticity in sex allocation, developmental mortality of males and limited insemination capacity of males. In all three cases the number of males per brood is expected to increase with brood size. Sex ratio optima may also be less female biased when several mothers contribute offspring to local mating groups or if non‐local mating occurs between members of different broods; again more males are expected in larger broods. In the parasitoid wasp Goniozus legneri (Hymenoptera: Bethylidae), sex allocation has only a small stochastic component, developmental mortality is low and non‐siblings are unlikely to develop in the same brood. However, the number of males per brood increases with the size of the brood (produced by a single mother). We investigated the further possibilities of limited insemination capacity and non‐local mating using a naturalistic experimental protocol. We found that limited insemination capacity is an unlikely general explanation for the increase in number of males with brood size. All males and females dispersed from both mixed and single sex broods. Although most females in mixed sex broods mated prior to dispersal, these data suggest that non‐local mating is possible, for instance via male immigration to broods containing virgin females. This may influence sex ratio optima and account for the trend in male number.
Call Intensity and Female Preferences in the European Green Toad
Authors: Castellano Sergio , Rosso Alessandra , Laoretti Francesca , Doglio Stefano , Giacoma Cristina 
Published: March 2003

AbstractIn the present paper we study the pattern of variation in call intensity in a natural population of the European green toad (Bufo viridis), and we analyse females preferences for this property by means of playback experiments. Although call sound pressure level (SPL) shows little within‐bout variation, we found significant positive correlation between call SPL and fundamental frequency: on average, an increase of 6 dB SPL produces a 100‐Hz increase in fundamental frequency. When females were given a choice between two calls differing by 10 and 6 dB SPL, they significantly preferred the loudest call, whereas they did not discriminate between calls differing by 3 dB. To test whether a 3‐dB difference reflects a sensory or behavioural limit, we carried out three experiments where females were given a choice between two calls differing in both duration (4 s and 6 s, respectively) and SPL (3 dB). In all three experiments, the longer call attracted a larger number of females, but the 3‐dB difference did not show any significant effect. Finally, we investigated the relationships between call intensity and frequency on female preferences. Previous experiments showed size‐dependent preferences for calls with lower‐than‐average frequencies (1.3 kHz) over calls with higher‐than‐average frequencies (1.6 kHz). Here, we show that this weak preference is abruptly reversed when the highest‐pitched call is broadcast at an intensity 6 dB higher than the alternative.
Adult Female Canaries Respond to Male Song by Calling
Authors: Nagle Laurent , Kreutzer Michel , Vallet Eric 
Published: March 2003

We describe a new assay for measuring the response to song playback by adult female domesticated canaries,Serinus canaria. We tested song perception and discrimination by measuring the frequency of particular female calls (`single calls') given in response to male song playbacks. We observed that females responded differently to songs of different species (canaries vs. pine siskin,Carduelis spinus) and to songs of different canary strains (domesticated vs. wild canaries). In addition, females were especially responsive to songs containing a particular type of song phrase (`A' phrases).This new assay provides equivalent results to the standard method (copulation solicitation displays) traditionally used to assess female song preferences. Our new assay has the advantage that it allows one to measure female song responsiveness without having to use estradiol implants and during both long and short day photoperiods. However, females responded differently in long and short days. We suggest that, by calling, females could both provide information about their sexual interest and attract the attention of particular males.