L-Amino acid oxidase from Naja naja oxiana venom

A new l-amino acid oxidase (LAAO) was isolated from the Central Asian cobra Naja naja oxiana venom by size exclusion, ion exchange and hydrophobic chromatography. The N-terminal sequence and the internal peptide sequences share high similarity with other snake venom l-amino acid oxidases, especially with those isolated from elapid venoms. The enzyme is stable at low temperatures (− 20 °C, − 70 °C) and loses its activity by heating at 70 °C. Specific substrates for the isolated protein are l-phenylalanine, l-tryptophan, l-methionine and l-leucine. The enzyme has antibacterial activity inhibiting the growth of Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) bacteria. N. naja oxiana LAAO dose-dependently inhibited ADP- or collagen-induced platelet aggregation with IC₅₀ of 0.094 μM and 0.036 μM, respectively. The antibacterial and anti-aggregating activity was abolished by catalase.     

The effect of short-term fasting and a single meal on protein synthesis and oxygen consumption in cod,Gadus morhua

Summary Rates of protein synthesis and oxygen consumption ( O₂) in cod were compared in both fasted and refed animals. During a 14-day fast both protein synthesis and respiration rates fell to stable values after 6 days. When a meal of whole sandeel at 6% body weight was fed to fish fasted for 6 days, protein synthesis and ( O₂) increased to a maximum at between 12 and 18 h after feeding. Peak ( O₂) was about twice the pre-feeding values, while whole animal protein synthesis increased four-fold. There were differences between tissues in the timing of maximum protein synthesis; the liver and stomach responded faster than the remainder of the body. Maximum protein synthesis rates in the liver and stomach occurred at 6 h after feeding, at which time their calculated contribution to total ( O₂) was 11%. Similar calculations suggested that the integrated increment in whole animal protein synthesis contributed between 23% and 44% of the post-prandial increase in ( O₂). It was concluded that protein synthesis is an important contributor to increased ( O₂) after feeding in cod.Abbreviations A _s absolute rate of protein synthesis - ASDA apparent specific dynamic action - ATP adenosine triphosphate - k _s fractional rate of protein synthesis - k _s/RNA amount of protein synthesized per unit RNA - ( O₂) oxygen consumption - PCA perchloric acid - RNA ribonucleic acid     

Trypsin from the pyloric caeca of bluefish (Pomatomus saltatrix)

Trypsin was purified from the pyloric caeca of bluefish (Pomatomus saltatrix) by ammonium sulfate precipitation, acetone precipitation and soybean trypsin inhibitor-Sepharose 4B affinity chromatography. Bluefish trypsin migrated as a single band using both sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and native-PAGE and had a molecular mass of 28 kDa. The optima pH and temperature for the hydrolysis of benzoyl-dl-arginine-p-nitroanilide (BAPNA) were 9.5 and 55 °C, respectively. The enzyme was stable over a broad pH range (7 to 12), but was unstable at acidic pH, and at temperatures greater than 40 °C. The enzyme was inhibited by specific trypsin inhibitors: soybean trypsin inhibitor (SBTI), N-p-tosyl-l-lysine chloromethyl ketone (TLCK) and the serine protease inhibitor phenylmethyl sulfonylfluoride (PMSF). CaCl₂ partially protected trypsin against activity loss at 40 °C, but NaCl (0 to 30%) decreased the activity in a concentration dependent manner. The N-terminal amino acid sequence of trypsin was determined as IVGGYECKPKSAPVQVSLNL and was highly homologous to other known vertebrate trypsins.     

Relationship between the octanol-water partition coefficient of tertiary amines and their effect of ‘selective’ uncoupling of photophosphorylation

A series of tertiary amines was investigated for effects on the transmembrane proton potential difference ( H), on photophosphorylation and on electron-flux control related to the intrathylakoid proton potential ( H_I), using isolated chloroplasts ofSpinacia oleracea L. As indicated by 9-aminoacridine fluorescence and [14C]methylamine uptake, all amines studied inhibited a build-up of H and, in parallel, ATP synthesis. Even when H was low, strong H₁-dependent electron-flux control was observed under the influence of tertiary amines. The strength of flux control in the presence of low H and the effectiveness of inhibition of ATP synthesis linearly increased with the lipophilicity of the amines. The most effective of the amines tested caused 50% inhibition of ATP synthesis at a concentration of 6 M, which is about 1000-fold lower than the concentration required for inhibition by methylamine. The data presented indicate the existence of two proton domains in the thylakoid vesicles, one of them feeding the ATP-synthase, the other the sites of pH-dependent electron-flux control. It is concluded that tertiary amines develop their action in a lipophilic domain of the thylakoid membrane, in the vicinity of the ATP-synthase complex. A mechanism for selective uncoupling and for the maintenance of H_I-dependent electron flux control in the presence of low H is discussed.Abbreviations and symbols coefficient for pH-dependent electron flux control - 9-AA 9-aminoacridine - Chl chlorophyll - I₅₀ amine concentration producing 50% inhibition of ATP-synthesis - J_e flux of photosynthetic electron transport - k _H apparent rate constant for proton efflux - H₁ proton potential in the thylakoid lumen - H₁ transthylakoid proton potential difference - p partition coefficient - q _AA coefficient for 9-aminoacridine fluorescence quenching - PS photosystem - Q quantum flux of photosynthetically active light Dedicated to Professor Wilhelm Simonis, on the occasion of his 80th birthday     

Effects of ambient temperature and altitude on ventilation and gas exchange in deer mice (Peromyscus maniculatus)

Summary The effects of different ambient temperatures (T _a) on gas exchange and ventilation in deer mice (Peromyscus maniculatus) were determined after acclimation to low and high altitude (340 and 3,800 m).At both low and high altitude, oxygen consumption ( ) decreased with increasingT _a atT _a from –10 to 30 °C. The was 15–20% smaller at high altitude than at low altitude atT _a below 30 °C.Increased atT _a below thermoneutrality was supported by increased minute volume ( ) at both low and high altitude. At mostT _a, the change in was primarily a function of changing respiration frequency (f); relatively little change occurred in tidal volume (V _T) or oxygen extraction efficiency (O₂EE). AtT _a=0 °C and below at high altitude, was constant due to decliningV _T and O₂EE increased in order to maintain high .At high altitude, (BTP) was 30–40% higher at a givenT _a than at low altitude, except atT _a below 10 °C. The increased at high altitude was due primarily to a proportional increase inf, which attained mean values of 450–500 breaths/min atT _a below 0 °C. The (STP) was equivalent at high and low altitude atT _a of 10 °C and above. At lowerT _a, (STPD) was larger at low altitude.At both altitudes, respiratory heat loss was a small fraction (<10%) of metabolic heat production, except at highT _a (20–30 °C).Abbreviations EHL evaporative heat loss - f respiration frequency - HL _a heat loss from warming tidal air - HL _e evaporative heat loss in tidal air - HL total respiratory heat loss - MHP metabolic heat production - O ₂ EE oxygen extraction efficiency - RQ respiratory quotient - T _a ambient temperature - T _b body temperatureT _lc lower critical temperature - carbon dioxide production - evaporative water loss - oxygen consumption - minute volume - V _T tidal volume     

Purification and characterisation of the cardenolide-specificβ-glucohydrolase CGH II from Digitalis lanata leaves

A cardenolide-hydrolysing β-D-glucosidase was isolated from young leaves of Digitalis lanata. Since this enzyme differs from the cardenolide glucohydrolase (CGH) described and characterised previously, it was termed cardenolide glucohydrolase II (CGH II). CGH II was detected in various Digitalis tissue cultures as well as in young leaves of D. lanata. The latter source was used as the starting material for the isolation and purification of CGH II. The specific enzyme activity reached about 15 pkat·mg^–1 protein in buffered leaf extracts. Optimal CGH II activity was seen at around pH 6.0 and 50 °C. CGH II was purified about 600-fold by anion exchange chromatography, size exclusion chromatography and hydroxyapatite chromatography. The apparent molecular mass of CGH II was 65 kDa as determined by SDS-PAGE. CGH II exhibited a high substrate specificity towards cardenolide disaccharides, especially to those with a 1-4-β-linked glucose-digitoxose moiety such as glucoevatromonoside. The K_m- and V_max-values for this particular substrate were calculated to be 101 μM and 19.8 nkat·mg^–1 protein, respectively.     

Thermoregulation,gas exchange,and ventilation in Adelie penguins (Pygoscelis adeliae)

Summary Adelie penguins (Pygoscelis adeliae) experience a wide range of ambient temperatures (T _a) in their natural habitat. We examined body temperature (T _b), oxygen consumption ( ), carbon dioxide production ( ), evaporative water loss ( ), and ventilation atT _a from –20 to 30 °C. Body temperature did not change significantly between –20 and 20°C (meanT _b=39.3°C).T _b increased slightly to 40.1 °C atT _a=30°C. Both and were constant and minimal atT _a between –10 and 20°C, with only minor increases at –20 and 30°C. The minimal of adult penguins (mean mass 4.007 kg) was 0.0112 ml/[g·min], equivalent to a metabolic heat production (MHP) of 14.9 Watt. The respiratory exchange ratio was approximately 0.7 at allT _a. Values of were low at lowT _a, but increased to 0.21 g/min at 30°C, equivalent to 0.3% of body mass/h. Dry conductance increased 3.5-fold between –20 and 30°C. Evaporative heat loss (EHL) comprised about 5% of MHP at lowT _a, rising to 47% of MHP atT _a=30°C. The means of ventilation parameters (tidal volume [VT], respiration frequency [f], minute volume [I], and oxygen extraction [ ]) were fairly stable between –20 and 10°C (VT did not change significantly over the entireT _a range). However, there was considerable inter- and intra-individual variation in ventilation patterns. AtT _a=20–30°C,f increased 7-fold over the minimal value of 7.6 breaths/min, and I showed a similar change. fell from 28–35% at lowT _a to 6% atT _a=30°C.Abbreviations C thermal conductance - EHL evaporative heat loss - oxygen extraction - f respiratory frequency - MHP metabolic heat production - evaporative water loss - LCT lower critical temperature - RE respiratory exchange ratio - T _a ambient temperature - T _b body temperature - rate of oxygen consumption - rate of carbon dioxide production - I inspiratory minute volume - VT tidal volume     

Effect of exposure to oxygen at 101 and 150 kPa on the cerebral circulation and oxygen supply in conscious rats

Hyperbaric oxygen at pressures of 300 to 500 kPa has been shown to induce changed distribution of cerebral blood flow ( _CBF) in rats, in places reducing the supply of the supplementary O₂. Thus, in the present study, the effect of hyperoxia at 101 (group 1, n = 9) and 150 (group 2, n = 9) kPa OZ on cerebral blood flow distribution and central haemodynamics was tested in conscious, habituated rats. During the control period the systolic arterial pressure (BP_s), heart rate (f _c), breathing frequency (f _b), cardiac output ( _c), arterial acid-base chemistry and glucose, as well as _CBF distribution (r _CBF) were similar in the two groups of animals. During O₂ exposure, the acid-base chemistry remained unchanged. The haemoglobin decreased in group 2, but remained unchanged in group 1. The f _c decreased rapidly in both groups during the change in gas composition, after which f _c remained constant both in group 1 and in group 2, for whom pressure was increased. The _c and f _b decreased and BP_s increased similarly in the two groups. Total _CBF and r _CBF decreased to the same extent in both groups, and the r _CBF changes were equally scattered. In group 1, breathing of pure O₂ did not increase the O₂ supply to any cerebral region except to the thalamus and colliculi after 60 min, whereas the O₂ supply to the hypothalamus decreased and remained low. In group 2, the O₂ supply was unchanged compared to the control period in all regions. These findings agree with previous observations during exposures to higher O₂ pressures. In air after O₂ exposure the acid-base chemistry remained normal. The f _c and f _b increased to higher levels than during the control period. The BP_s remained high. The brain blood flows were increased, inducing elevated O₂ supply to several brain regions compared to the control period. In conclusion, O₂ supply to the central nervous system was found to be in the main unchanged during breathing of O₂ at 101 kPa and 150 kPa.     

Thermoregulation in a large bird,the emu (Dromaius novaehollandiae)

The emu is a large, flightless bird native to Australia. Its habitats range from the high snow country to the arid interior of the continent. Our experiments show that the emu maintains a constant body temperature within the ambient temperature range-5 to 45°C. The males regulate their body temperature about 0.5°C lower than the females. With falling ambient temperature the emu regulates its body temperature initially by reducing conductance and then by increasing heat production. At-5°C the cost of maintaining thermal balance is 2.6 times basal metabolic rate. By sitting down and reducing heat loss from the legs the cost of homeothermy at-5°C is reduced to 1.5 times basal metabolic rate. At high ambient temperatures the emu utilises cutaneous evaporative water loss in addition to panting. At 45°C evaporation is equal to 160% of heat production. Panting accounts for 70% of total evaporation at 45°C. The cost of utilising cutaneous evaporation for the other 30% appears to be an increase in dry conductance.Abbreviations A _r Effective radiating surface area - BMR basal metabolic rate - C _dry dry conductance - CEWL cutaneous evaporative water loss - EHL evaporative heat loss - EWL evaporative water loss - FECO₂ fractional concentration of CO₂ in excurrent air - FF_H2O water content of chamber excurrent air - FEO₂ fractional concentration of O₂ in chamber excurrent air - FICO₂ fractional concentration of CO₂ in incurrent air - FIO₂ fractional concentration of O₂ in chamber incurrent air - MHP metabolic heat production - MR metabolic rate - REWL respiratory evaporative water loss - RH relative humidity - RQ respiratory quotient ; - SA surface area - SEM standard error of the mean - SNK Student-Newman-Keuls multiple range test - STPD standard temperature and pressure dry - T _a ambient temperature(s) - T _b body temperature(s) - T _e surface temperature(s) - flow rate of air into the chamber - carbon dioxide production - oxygen consumption - vapour pressure of water     

Characterization of phenoloxidase activity in Sydney rock oysters (Saccostrea glomerata)

Phenoloxidase (PO) activity was studied in Sydney rock oysters (Saccostrea glomerata). As in other molluscs, PO was found to exist as a pro-enzyme (proPO) in hemocytes. ProPO could be activated to PO by exogenous proteases (trypsin and chymotrypsin), exposure of hemocytes to pathogen-associated molecular patterns (PAMPs) and by the detergents, Triton X-100 and sodium dodecyl sulphate (SDS). Inhibition studies confirmed the proPO activating system of Sydney rock oysters is a proteinase cascade in which Ca²⁺ dependent serine proteinases proteolytically convert proPO into active PO. Activated PO was found to be a tyrosinase-like enzyme that is responsible for both monophenolase and diphenolase activity. The bifunctional PO had higher affinity for the monophenol, hydroquinine monomethyl ether (4HA) (K_m = 4.45 ± 1.46 mM) than for the diphenol, l-DOPA (K_m = 10.27 ± 1.33 mM). Maximum enzyme activity was evident at 37 °C, pH 8 and at salinities of between 30 and 37 ppt. Melanogenesis catalysed by the active enzyme is a composite of eumelanin and the product of a sclerotin pathway combining DOPA decarboxylase with PO activity.     

Cardiovascular responses to facial cooling during low and moderate intensity exercise

To investigate the hypothesis that facial cooling (FC) exerts a greater influence on the cardiovascular system at lower versus higher levels of exercise, this study examined the effect of facial cooling [mean (SE): 0 (2)°C at 0.8 m·s^–1 wind velocity] during 30 min low [35% maximum oxygen consumption ( O_2max)] and moderate (70% O_2max) levels of cycle ergometry in the supine position. Five male subjects were assigned in random order to four exercise conditions: (1) FC at 35% O_2max(FC35), (2) no cooling (NFC35), (3) FC at 70% O_2max(FC70), and (4) no cooling (NFC70). Heart rate (f _c), stroke volume (V _s), and cardiac output ( _c) were measured at rest and every 10 min of exercise using impedance cardiography. During FC35, the change in f _c [mean (SE)] was significantly lower (P < 0.05) than NFC35 at 10 [22 (5) vs 31 (3) beats· min^–1], 20 [29 (6) vs 35 (3) beats·min^–1], and 30 [29 (5) vs 38 (4) beats·min^–1] min. No differences in f _c were observed between FC70 and NFC70. Furthermore, FC had no effect on V _s or _cat either exercise intensity. However, when comparing the FC70 and NFC70 conditions, there was a significant main effect (P<0.05) in mean arterial pressure (P _a) response with cooling despite the fact that neither V _s or _cwere different from the NFC70 control. The increase (P < 0.05) in the estimated change in systemic vascular resistance ( _a· _c ^–1) could partly explain the relative rise in _aat FC70. No pressor effect of cooling was observed at 35% O_2max. The results suggest that the FC condition promotes exercise bradycardia at low levels of exercise and exerts a greater pressor response during moderate exercise.     

A method for determining the maximal steady state of blood lactate concentration from two levels of submaximal exercise

The aim of this study was to estimate the characteristic exercise intensity _CL which produces the maximal steady state of blood lactate concentration (MLSS) from submaximal intensities of 20 min carried out on the same day and separated by 40 min. Ten fit male adults [maximal oxygen uptake _max 62 (SD 7) ml · min^–1 · kg^–1] exercisOed for two 30-min periods on a cycle ergometer at 67% (test 1.1) and 82% of _max (test 1.2) separated by 40 min. They exercised 4 days later for 30 min at 82% of _max without prior exercise (test 2). Blood lactate was collected for determination of lactic acid concentration every 5 min and heart rate and O₂ uptake were measured every 30 s. There were no significant differences at the 5th, 10th, 15th, 20th, 25th, or 30th min between , lactacidaemia, and heart rate during tests 1.2 and 2. Moreover, we compared the exercise intensities _CL which produced the MLSS obtained during tests 1.1 and 1.2 or during tests 1.1 and 2 calculated from differential values of lactic acid blood concentration ([1a^–]_b) between the 30th and the 5th min or between the 20th and the 5th min. There was no significant difference between the different values of _CL [68 (SD 9), 71 (SD 7), 73 (SD 6),71 (SD 11) % of _max (ANOVA test,P<0.05). Four subjects ran for 60 min at their _CL determined from periods performed on the same day (test 1.1 and 1.2) and the difference between the [la^–]_b at 5 min and at 20 min ( ([la^–]_b)) was computed. The [la^–]_b remained constant during exercise and ranged from 2.2 to 6.7 mmol · l^–1 [mean value equal to 3.9 (SD 1) mmol · l^–1]. These data suggest that the _CL protocol did not overestimate the exercise intensity corresponding to the maximal fractional utilization of _max at MLSS. For half of the subjects the _CL was very close to the higher stage (82% of _max where an accumulation of lactate in the blood with time was observed. It can be hypothesized that _CL was very close to the real MLSS considering the level of accuracy of [la^–]_b measurement. This study showed that exercise at only two intensities, performed at 65% and 80% of _max and separated by 40 min of complete rest, can be used to determine the intensity yielding a steady state of [la^–1]_b near the real MLSS workload value.     

Characterization of arylalkylamine N-acetyltransferase (AANAT) activities and action spectrum for suppression in the band-legged cricket, Dianemobius nigrofasciatus (Orthoptera: Gryllidae)

Arylalkylamine N-acetyltransferase (AANAT), constituting a large family of enzymes, catalyzes the transacetylation from acetyl-CoA to monoamine substrates, although homology among species is not very high. AANAT in vertebrates is photosensitive and mediates circadian regulation. Here, we analyzed AANAT of the cricket, Dianemobius nigrofasciatus. The central nervous system contained AANAT activity. The optimum pHs were 6.0 (a minor peak) and 10.5 (a major peak) with crude enzyme solution. We analyzed the kinetics at pH 10.5 using the sample containing collective AANAT activities, which we term AANAT. Lineweaver-Burk plot and secondary plot yielded a K_m for tryptamine as substrate of 0.42 µM, and a V_max of 9.39 nmol/mg protein/min. The apparent K_m for acetyl-CoA was 59.9 µM and the V_max was 8.14 nmol/mg protein/min. AANAT of D. nigrofasciatus was light-sensitive. The activity was higher at night-time than at day-time as in vertebrates. To investigate most effective wavelengths on AANAT activity, a series of monochromatic lights was applied (350, 400, 450, 500, 550, 600 and 650 nm). AANAT showed the highest sensitivity to around 450 nm and 550 nm. 450 nm light was more effective than 550 nm light. Therefore, the most effective light affecting AANAT activity is blue light, which corresponds to the absorption spectrum of blue wave (BW)-opsin.     

Physiological responses to cold (10° C) in men after six months' practice of yoga exercises

A study was conducted on 30 healthy soldiers (age: 40–46 years) to assess the effect of selected yogic exercises (asanas) on some physiological responses to cold exposure. They were randomly divided into two groups of 15 each. One group performed regular physical exercises of physical training (PT), while the other group practised yogic exercises. At the end of 6 months of training, both the groups were exposed together to cold stress at 10°C for 2 h, and the following parameters were periodically monitored during cold exposure: heart rate (fH), blood pressure (BP), cardiac output , oral temperature (T_or), skin temperature (T _sk), respiratory rate (fR), minute ventilation , oxygen consumption , and shivering response by integrated electromyogram (EMG). There were progressive increases inBP, fR, , , and and decreases infH,T _or andT _sk during cold exposure in both the groups. However, the decrease inT _or and the increases in and were relatively lower (P<0.01) in the yoga group as compared to the PT group. The shivering response appeared much earlier and was more intense in the PT group. These findings suggest that practice of yoga exercises may improve cold tolerance.     

Growth and protein synthesis of barramundi, Lates calcarifer, fed lupin as a partial protein replacement

Protein synthesis is an essential growth process in all animals. Little information is available on post-prandial protein synthesis and even less where different protein sources are compared. Protein synthesis was measured at 4 and 24 h after feeding juvenile barramundi in order to determine the effect of using lupin as a partial protein replacement for fish meal on the post-prandial protein metabolism. Juvenile barramundi (4.3 ±0.6 g) were held in a recirculation system (27 °C, salinity 10‰ and 24 h light) for 15 days. Fish were fed one of two isonitrogenous isoenergetic diets (40% crude protein, 16% lipid and 18.5 GE MJ kg^− 1). One diet was formulated with 100% fish meal as the protein source while the other had 45% of the protein replaced with lupin ingredients (lupin kernel meal (Lupinus angustifolius) and lupin protein concentrate). All fish were fed a ration of 6%·d^− 1 and feed intake was not significantly different between the two diets. Specific growth rate (SGR) and growth efficiency (in relation to protein (PPV) and energy (PEV)) were 6.5 ± 0.14%·d^− 1, 43.8 ± 2.72% and 38.31 ± 1.56%, respectively, and were not significantly different between the two diets. There was no significant difference in protein synthesis between the two diets at 4 and 24 h after feeding, however protein synthesis was significantly higher 4 h after feeding than at 24 h (p = 0.02). Neither growth performance nor protein metabolism was altered by replacing 45% of the protein with lupin protein and indicated this to be a suitable protein source for barramundi feeds.     

Mechanisms of potentiation in sweating induced by long-term physical training

To evaluate the mechanism of potentiation of sweating after long-term physical training, we compared sweating function in trained and untrained subjects using the frequency of sweat expulsion (f _sw) as an indicator of central sudomotor activity. Nine trained male subjects (trained group) and eight untrained male subjects (untrained group) performed 30-min cycle exercise at 35% maximal oxygen uptake at 25°C ambient temperature and 35% relative humidity. Oesophageal temperature (T _oes), mean body temperature _b, chest sweating rate ( _sw,chest), forearm sweating rate ( _forearm), andf _sw were measured. The slopes of the _sw,chest versus body temperature (T _oes and _b) and versusf _sw relationships in the trained group were significantly greater than those in the untrained group (both,P < 0.05), while there was no difference between the groups in the slopes of the _sw,chest versus body temperature or versusf _sw relationships. Neither the body temperature threshold for initiation of chest or forearm sweating nor the slope of thef _sw- _b relationship differed between groups. We concluded that, during light exercise at moderate ambient temperature, the _sw,chest in the subjects who had undergone long-term physical training was greater than that in the untrained subjects while the _sw,forearm was not changed. The greater _sw,chest in the trained subjects was concluded to be due to an increase of sensitivity of peripheral mechanisms.     

Molecular cloning and characterization of the gene encoding a fibrinolytic enzyme from <Emphasis Type="Italic">Bacillus subtilis</Emphasis> Strain A1

A fibrinolytic metalloprotease gene from Bacillus subtilis has been cloned in Escheridria coliXL1-Blue and the bacterial expressed enzyme was purified. The nucleotide sequence of the cloned fibrinolytic enzyme gene revealed a single open reading frame of 1023 bp coding for 341 amino acids (M _r 37708.21 Da). N-terminal amino acid sequencing of the fibrinolytic enzyme excreted from E. coli host cells revealed that the mature fibrinolytic enzyme consists of 288 amino acids (M _r 31391.1 Da). The deduced amino acid sequence showed significant homology with Erwina carotovora neutral metalloprotease and Serratia marcescens minor metalloprotease by 65 and 58% amino acid sequence identity, respectively. The protein showed significant alignments with the conserved domain of catalytic activity and the -helix domain in Bacillus anthracisthermolysis metalloprotease. The biochemical properties of the purified enzyme suggested that the enzyme is a fibrinolytic metalloprotease, which has optimal activity at pH 7.0 and 50 °C.     

Evaluation of physical fitness from field tests at high altitude in circumpubertal boys: comparison with laboratory data

Field tests of running and laboratory tests were performed in La Paz [high altitude (HA), 3700 m] and in Clermont-Ferrand [low altitude (LA), 300 m] to investigate their validity at HA. Prepubertal boys of mean ages 10.6 years (HA1,n = 16; LA1,n = 28) and pubertal boys of 13.7 years (HA2,n = 12; LA2,n = 41) took part in the study. All the boys performed a 30-m sprint (v _30m), a 30-s shuttle run (v _3os) and a progressive shuttle run test until their maximal aerobic velocity (v _maxsRT). Maximal oxygen consumption was extrapolated from the last test. . In the laboratory, the boys performed a force-velocity test (P _max), a Wingate test (P _Wing) and a graded test to measure maximal oxygen consumption ; direct method) on a cycle ergometer. At similar ages, there was no significant difference between HA and LA boys forv _30m andP _max. Thev _30s of HA boys was 3%–4% lower than those of LA boys (P<0.05); there was no significant difference forP _Wing. Significant relationships were observed at both altitudes betweenP _max (watts per kilogram) andv _30m (HA:r=0.76; LA:r=0.84) and betweenP _Wing andv _30s (HA:r=0.67; LA:r = 0.77); the slopes and the origins were the same at HA and LA. The ,v _maxSRT and were lower by 9%, 12% and 20%, respectively, at HA than at LA (P<0.05). However, the relationships between and (litres per minute) at HA (r=0.88) and at LA (r=0.93) were identical. In conclusion, chronic hypoxia did not modify performance in very short dash exercises. The influence of HA appeared when the exercise duration increased and, during a maximal shuttle run test, performance was reduced by 10% at HA. Moreover, it was possible to assessP _max,P _Wing and at HA as well as at LA from field tests.