Sublimation in bright spots on (1) Ceres.

The dwarf planet (1) Ceres, the largest object in the main asteroid belt with a mean diameter of about 950 kilometres, is located at a mean distance from the Sun of about 2.8 astronomical units (one astronomical unit is the Earth-Sun distance). Thermal evolution models suggest that it is a differentiated body with potential geological activity. Unlike on the icy satellites of Jupiter and Saturn, where tidal forces are responsible for spewing briny water into space, no tidal forces are acting on Ceres. In the absence of such forces, most objects in the main asteroid belt are expected to be geologically inert. The recent discovery of water vapour absorption near Ceres and previous detection of bound water and OH near and on Ceres (refs 5-7) have raised interest in the possible presence of surface ice. Here we report the presence of localized bright areas on Ceres from an orbiting imager. These unusual areas are consistent with hydrated magnesium sulfates mixed with dark background material, although other compositions are possible. Of particular interest is a bright pit on the floor of crater Occator that exhibits probable sublimation of water ice, producing haze clouds inside the crater that appear and disappear with a diurnal rhythm. Slow-moving condensed-ice or dust particles may explain this haze. We conclude that Ceres must have accreted material from beyond the 'snow line', which is the distance from the Sun at which water molecules condense.

Self-rated health as a predictor of outcomes of type 2 diabetes patient education programmes in Denmark.

OBJECTIVE: To explore if self-rated health (SRH) can predict differences in outcomes of patient education programmes among patients with type 2 diabetes over time. STUDY DESIGN: This is an observational cohort study conducted among 83 patients with type 2 diabetes participating in patient education programmes in the Capital Region of Denmark. METHODS: Questionnaire data were collected by telephone interview at baseline and 2 weeks (77 participants, 93%) and 12 months (66, 80%) after the patient education ended. The seven-scale Health Education Impact Questionnaire (HeiQ) was the primary outcome. The independent variable was SRH, which was dichotomized into optimal or poor SRH. Changes over time were assessed using mean values and standard deviation (SD) at each time point and Cohen effect sizes. Odds ratios and 95% confidence intervals were calculated for the likelihood of having poor SRH for each baseline sociodemographic and health-related variable. RESULTS: Twelve months after patient education programmes, 60 (72%) patients with optimal SRH at baseline demonstrated increased self-management skills, overall acceptance of chronic illness, positive social interaction with others, and improved emotional well-being. Participants with poor SRH (23, 28%) reported no improvements over time. Not being married (odds ratio [OR] 7.79, P < 0.001), living alone (OR 4.93, P = 0.003), having hypertension (OR 8.00, P = 0.031), and being severely obese (OR 4.07, P = 0.009) were significantly associated with having poor SRH. After adjusting for sex, age and vocational training, marital status (OR 9.35, P < 0.001), cohabitation status (OR = 4.96, P = 0.005) and hypertension (OR 10.9, P = 0.03) remained associated with poor SRH. CONCLUSIONS: We found a strong association between SRH and outcomes of patient education, as measured by the HeiQ, at 12 months. Only participants with optimal SRH appeared to benefit from patient education. Other patient characteristics may be responsible to explain the observed difference between patients with optimal and poor SRH.

Association of physical workload and leisure time physical activity with incident mobility limitations: a follow-up study.

OBJECTIVES: To examine individual as well as joint associations of physical workload and leisure time physical activity with incident mobility limitations in initially well-functioning middle-aged workers. METHODS: This study is based on 6-year follow-up data of the Danish Longitudinal Study on Work, Unemployment and Health. Physical workload was reported at baseline and categorised as light, moderate or heavy. Baseline leisure time physical activity level was categorised as sedentary or active following the current recommendations on physical activity. Incidence of mobility limitations in climbing stairs and running among initially well-functioning workers (n=3202 and n=2821, respectively) was assessed during follow-up. RESULTS: Higher workload increased whereas active leisure time decreased the risk of developing mobility limitations. The incidence of limitations increased progressively with higher workload regardless of level of leisure time physical activity, although the risks tended to be higher among those with sedentary leisure time compared with their active counterparts. All in all, the risk for onset of mobility limitations was highest among those with heavy workload combined with sedentary leisure time and lowest among those with light workload combined with active leisure time. CONCLUSIONS: Although leisure time physical activity prevents development of mobility decline, high workload seems to accelerate the progression of mobility limitations among both those with active and sedentary leisure time. Therefore, efforts should be made to recommend people to engage in physical activity regardless of their physical workload.

How do young men want to receive information about fertility? Young men's attitudes towards a fertility campaign targeting men in Copenhagen, Denmark.

STUDY QUESTION: What are young men's attitudes towards the Danish fertility campaign 'How's your sperm?' and how do they want to receive fertility information in the future? SUMMARY ANSWER: The young men found that the campaign had limited impact because it was not relevant to their current life situation and they believed general fertility awareness should be a mandatory part of education, while more targeted information would be helpful through web-based venues when it was more relevant to their lives (e.g. when ready to have children). WHAT IS KNOWN ALREADY: It is estimated that 16-26% of the Danish population who want children will experience infertility at some point in their lives. In Denmark, 25% of young healthy men have decreased sperm quality, and 20% of 50-year-old men are childless. Men play an important role in the fertility decision-making of couples, thus, it is important to target men and ensure that they have sufficient fertility knowledge. However, fertility awareness is limited among men and there have been few fertility awareness initiatives targeting men. In October 2018, the Municipality of Copenhagen launched the campaign 'How's your sperm?' as a tool to increase fertility knowledge among men. To identify potential barriers for the effect of fertility campaigns targeting men, evaluations of such campaigns are needed. STUDY DESIGN SIZE DURATION: This study was a cross-sectional, qualitative study of six focus groups including a total of 27 currently childless young men from the Capital Region of Copenhagen, Denmark. Data collection took place between April and October 2019. PARTICIPANTS/MATERIALS SETTING METHODS: The interviewed young men were currently childless and were all residents in the Capital Region of Copenhagen. They were between 23 and 32 years old with an average age of 26 years, and almost all were university students or had a university degree. The focus group discussions were audiotaped, anonymized and transcribed in full. Data were analyzed using qualitative content analysis. MAIN RESULTS AND THE ROLE OF CHANCE: Overall, the campaign had limited influence on the young men because they believed the campaign was not relevant to their current life situation. Furthermore, the young men were confused about the aim and message of the campaign, as they thought it encouraged them to have their sperm quality tested. The young men also criticized the campaign for making a link between sperm quality and masculinity. They recognized the importance of knowledge about reproductive health but they wanted access to accurate information about fertility and risk factors for infertility. According to the young men, future initiatives should prioritize clear communication of accurate, reliable and understandable fertility information in web-based venues. In addition, the young men suggested that general fertility information should be a mandatory part of the (sexual) education curriculum in primary and secondary schools. LIMITATIONS REASONS FOR CAUTIONS: Participants were young and highly educated; thus the findings cannot be generalized to all men of a similar age group or to men at older ages. WIDER IMPLICATIONS OF THE FINDINGS: Different strategies that are relevant to the lifespan are needed to increase fertility awareness in the male population. The young men's responses underscored that any fertility awareness strategy or campaign must convey respect for the individual's autonomy. The findings highlight that how information is communicated and the quality or type of information that is disseminated are both important in acceptability by target users. STUDY FUNDING/COMPETING INTERESTS: No funding was received for this study. No conflicts were declared for all authors. TRIAL REGISTRATION NUMBER: N/A.

The impact of social relations on the incidence of severe depressive symptoms among infertile women and men.

BACKGROUND: The present study investigates anticipated support and perceived appreciation as well as perceived negative aspects of social relations as determinants of the incidence of severe depressive symptoms among women and men in unsuccessful fertility treatment. METHODS: A prospective cohort study of consecutively recruited, new couples in fertility treatment completed questionnaires immediately before initiating treatment (T1) and at 1-year follow-up (T2) (response rates 80%, 88%). The study population included 695 participants (355 women and 340 men) who had no severe depressive symptoms at T1, who had no child together at T1 and whose treatment had failed. We measured severe depressive symptoms with Mental Health Inventory 5 from Short-Form 36, functional aspects of general social relations and of infertility-specific social relations. RESULTS: Fifteen per cent of women and 6% of men in unsuccessful treatment reported severe depressive symptoms at T2. Among men, low emotional support, low appreciation and high excessive demands from the partner were significant determinants of the incidence of severe depressive symptoms. Among women and men, low appreciation from the family, many conflicts and high excessive demands from family, friends and neighbours were significant determinants of severe depressive symptoms. The analyses were controlled for gender. Duration of infertility, infertility diagnosis and socio-economic position did not confound the associations. CONCLUSIONS: More women than men report severe depressive symptoms after 1 year of unsuccessful treatment. It is important to be aware of the possible negative impact of relational strain among fertility patients in unsuccessful treatment.

The longitudinal impact of partner coping in couples following 5 years of unsuccessful fertility treatments.

BACKGROUND: Because there is a lack of longitudinal research examining the impact of partner coping in couples experiencing infertility, we know very little about the long-term nature of coping with infertility and how partner coping strategies impact personal, marital and social distress. METHODS: Participants were Danish men and women about to start a cycle of assisted reproduction treatment who were followed for a 5 year period of unsuccessful treatments. Multilevel modeling using the actor-partner interdependence model was used to examine the couple as the unit of analysis. RESULTS: Active and passive avoidance coping strategies were significantly related to increased personal, marital and social distress at the individual and partner level. Meaning-based coping strategies were related to decreases in a woman's individual distress and her partner's marital distress. CONCLUSIONS: Partner coping strategies have a significant impact on the other member of the couple over time in men and women undergoing infertility treatments over a 5 year period. Physicians and mental health professionals can educate men and women regarding the ineffectiveness of avoidance coping strategies as well as the beneficial nature of finding new meaning and life goals while experiencing the stress of infertility.

SEIS: Insight's Seismic Experiment for Internal Structure of Mars.

By the end of 2018, 42 years after the landing of the two Viking seismometers on Mars, InSight will deploy onto Mars' surface the SEIS (Seismic Experiment for Internal Structure) instrument; a six-axes seismometer equipped with both a long-period three-axes Very Broad Band (VBB) instrument and a three-axes short-period (SP) instrument. These six sensors will cover a broad range of the seismic bandwidth, from 0.01 Hz to 50 Hz, with possible extension to longer periods. Data will be transmitted in the form of three continuous VBB components at 2 sample per second (sps), an estimation of the short period energy content from the SP at 1 sps and a continuous compound VBB/SP vertical axis at 10 sps. The continuous streams will be augmented by requested event data with sample rates from 20 to 100 sps. SEIS will improve upon the existing resolution of Viking's Mars seismic monitoring by a factor of ∼ 2500 at 1 Hz and ∼ 200 000 at 0.1 Hz. An additional major improvement is that, contrary to Viking, the seismometers will be deployed via a robotic arm directly onto Mars' surface and will be protected against temperature and wind by highly efficient thermal and wind shielding. Based on existing knowledge of Mars, it is reasonable to infer a moment magnitude detection threshold of M w ∼ 3 at 40 ∘ epicentral distance and a potential to detect several tens of quakes and about five impacts per year. In this paper, we first describe the science goals of the experiment and the rationale used to define its requirements. We then provide a detailed description of the hardware, from the sensors to the deployment system and associated performance, including transfer functions of the seismic sensors and temperature sensors. We conclude by describing the experiment ground segment, including data processing services, outreach and education networks and provide a description of the format to be used for future data distribution. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s11214-018-0574-6) contains supplementary material, which is available to authorized users.

The impact of partner coping in couples experiencing infertility.

BACKGROUND: Most studies examining coping with infertility use the individual as the unit of analysis. Although valuable, these studies fail to show the impact that partner coping has on individual distress. Since infertility is a shared stressor, examining the impact of partner coping is particularly relevant. METHODS: Data were based on a questionnaire in a consecutive sample of 1169 women and 1081 Danish men prior to beginning assisted reproduction treatment. Multilevel modeling using the Actor Partner Interdependence Model and follow-up analysis of variance were used to examine the couple as the unit of analysis. RESULTS: A partner's use of active-avoidance coping was related to the increased personal, marital and social distress for men and women. A woman's use of active-confronting coping was related to increased male marital distress. And a partner's use of meaning-based coping was associated with decreased marital distress in men and increased social distress in women. CONCLUSIONS: Although understudied, partner coping patterns play a key role in a partner's ability to cope with the infertility experience. Physicians and mental health providers can help couples to understand the coping strategies that lead to increased and decreased partner distress.

Urokinase-catalysed plasminogen activation. Effects of ligands binding to the AH-site of plasminogen.

The kinetics of activation of Lys-plasminogen (Lys-77-Asn-790) and miniplasminogen (Val-442-Asn-790) catalysed by low-molecular-weight urokinase (LMW-urokinase) was investigated in the presence and absence of ligands that bind to the AH-site of the plasminogens. 6-Aminohexanoic acid and alpha-N-acetyl-L-lysine methyl ester (AcLysMe) were used. Saturation of the AH-sites of the plasminogens result in similar, but rather small positive effects on the kinetics of activation of the two plasminogens. Michaelis constants decrease approx. 2-fold and second-order rate constants (kc/Km)Pg increase approx. 1.2-fold. Michaelis constants (KPg values) were obtained using a new approach; the values were determined from the competing effects of the plasminogens on urokinase-catalysed hydrolysis of a synthetic substrate. In the pH range 7.4-8.0, only minor alterations of the values of the kinetic parameters are observed. At 25 degrees C, values of (kc/Km)Pg are approx. 3-fold less than the value at 37 degrees C, whereas KPg is not changed. We conclude that kc/Km values are approx. 10(5) M-1.s-1 and that KPg values are approx. 40 microM of urokinase-catalysed conversions of Lys- and miniplasminogen to their respective plasmins.

Studies on the nature of the catalytically essential ionizing group of plasmin with pK 8.4.

A fully carbamylated derivative of plasminogen having no free amino groups has been prepared and converted by urokinase to an active enzyme, called carbamyl plasmin A, with a single free NH2-terminal amino group (Val-561). Carbamyl plasmin A was shown to possess a catalytically essential ionizing group having pK 8.6. Carbamylation of the free NH2-terminal amino group of carbamyl plasmin A led to complete loss of catalytic activity. The results of solvent perturbation studies of normal plasmin (EC 3.4.21.7) indicate that the group with pK 8.4 is a neutral acid group. It is suggested that the catalytically essential ionizing group of plasmin having a pK of 8.4 is the alpha-ammonium group of the NH2-terminal Val-561 or the light chain of plasmin, forming an ion pair with a COO- group of an aspartate or glutamate residue.

Allosteric effects of some antifibrinolytic amino acids on the catalytic activity of human plasmin.

The effects of L-lysine, 6-aminohexanoic acid, and trans-4-aminomethylcy-clohexane-1-carboxylic acid on the catalytic activity of plasmin (EC 3.4.21.7) have been investigated. The kinetics of the plasmin-catlysed hydrolysis of alpha-N-benzoyl-L-arginine ethyl ester in the presence of these compounds have been studied at a number of different concentrations of the three modifiers. They each exert two effects on the reaction, an activation and an inhibition, the concentration dependencies of which are markedly different. They must therefore arise from two different interactions between plasmin and the modifier. The inhibition is competitive, so that it most probably results from direct interaction at the catalytic site. The activation is kinetically non-competitive. The experimental observations seem to be explained best by assuming that L-lysine and certain analogous compounds function as both allosteric modifiers and competitive inhibitors of plasmin.

pH effects in plasmin-catalysed hydrolysis of alpha-N-benzoyl-L-arginine compounds.

The steady-state kinetics of plasmin (EC 3.4.21.7) catalysed reactions with some alpha-N-benzoyl-L-arginine compounds is investigated in the pH range 5.8--9.0. The results are interpreted in terms of a three-step mechanism, which involves enzyme-substrate complex formation, followed by acylation and deacylation of the enzyme. Alpha-N-Benzoyl-L-arginine methyl ester and ethyl ester show the same pH behaviour. The kinetic parameter kc/Km is influenced by two groups with pK values of 6.5 and 8.4, respectively. kc is affected only by the group with pK equal to 6.5 and Km only by the group with pK equal to 8.4. It is suggested that the group with pK equal to 6.5 is the 1-chloro-3-tosyl-amido-7-amino-2-heptanone-sensitive histidine residue in the active site and that the group with pK equal to 8.4 is perhaps the alpha-amino group of the N-terminus in analogy to trypsin and chymotrypsin. alpha-N-Benzoyl-L-arginine amide is not hydrolysed by plasmin, but proves to be a competitive inhibitor, Ki = 12.8 +/- 1.8 mM, pH = 7.8. Also the product alpha-N-benzoyl-L-arginine is a competitive inhibitor, Ki = 26 +/- 3.1 mM, pH = 7.8. Estimates of individual rate constants are compared with similar trypsin data.

Kinetic studies of the urokinase-catalysed conversion of NH2-terminal glutamic acid plasminogen to plasmin.

Initial velocities for the urokinase (EC 3.4.99.26)-catalysed conversion of glutamic acid plasminogen to plasmin (EC 3.4.21.7) have been determined at various urokinase and glutamic acid plasminogen concentrations. As has been found for the corresponding reaction with lysine plasminogen this conversion obeys the Michaelis rate equation. The apparent Michaelis constants are of the same order of magnitude for lysine and glutamic acid plasminogens. The difference in conversion rates for the reactions has been shown to be connected with their having different catalytic constants. The data were analysed according to two reaction schemes, in one of which only one peptide bond is split during the glutamic acid plasminogen-plasmin conversion and in the other of which the cleavage of two peptide bonds with the obligatory formation of an intermediate plasminogen is assumed. The results favour the former.

Kinetic studies of the urokinase catalysed conversion of NH2-terminal lysine plasminogen to plasmin.

A method for determining initial velocities of the urokinase (EC 3.4.99.26) catalysed converstion of NH2-terminal lysine plasminogen to plasmin (EC 3.4.21.7) is presented. This reaction has been coupled with the hydrolysis of alpha-N-benzyoly-L-arginine ethyl ester, which is catalysed by plasmin, and its rate has been determined from the time course of the overall reaction. The proenzyme-enzyme conversion was found to obey the Michaelis-Menten rate equation. The following values of the kinetic parameters were obtained: the apparent Michaelis constant, Km = 40.7 +/- 6.2 muM; the catalytic constant, kc = 2.59+/-0.31 s(-1), and kc/Km = 6.36-10(4) +/- 0.24-10(4) M(-1)-s(-1).

Activity modulation of the fast and slow isozymes of human cytosolic low-molecular-weight acid phosphatase (ACP1) by purines.

The activity modulation of homogeneous isozymes of the human cytosolic M(r) 18,000 acid phosphatase (ACP1) by purines has been investigated. A pronounced difference in the response of fast and slow isozymes of the same genetic type was observed, while identical properties were found for fast isozymes encoded by different alleles (ACP1 X A, B and C), as well as for the corresponding slow isozymes. The catalytic rate constant (kc) of the fast isozymes was increased 5.1-fold by hypoxanthine and decreased 40% by adenine, while the kc of the slow isozymes was unaffected by hypoxanthine but increased 4.6-fold by adenine. This finding and the genetically-determined differences in the relative quantities of the fast and slow isozymes account for the well-known phenotypic differences in activity modulation. The kinetic results strongly indicate that the effector binds to the free enzyme, as well as to the enzyme-substrate complex. Activating effectors showed a higher affinity for the free enzyme than for the enzyme-substrate complex, while the reverse was true with the inhibitor. The results exclude the possibility that effector and substrate bind to the same site of the enzyme; parasteric binding to adjacent sites is suggested.

Steady-state kinetics of plasmin- and trypsin-catalysed hydrolysis of a number of tripeptide-p-nitroanilides.

The steady-state kinetics of plasmin- (EC 3.4.21.7) and trypsin-catalysed (EC 3.4.21.4) hydrolysis of Bz-L-Phe-Val-Arg-pNA, Bz-D-Phe-Val-Arg-pNA, L-Phe-Val-Arg-pNA, D-Phe-Val-Arg-pNA and D-Val-Leu-Lys-pNA were investigated in the pH range 6-9. The pH dependences of the kinetic parameters correspond with the effects of catalytically essential ionizations in the enzymes, except for reactions with L- and D-Phe-Val-Arg-pNA, in which protonation of the NH2-terminal alpha-amino groups (pK = 7.0) shows some inhibitory effect. The reactions of plasmin and trypsin with p-nitroanilides show kc values similar to those normally found with specific ester substrates, indicating that the deacylation steps of the reactions are rate determining.

Kinetic studies of urokinase-catalysed hydrolysis of 5-oxo-L-prolylglycyl-L-arginine 4-nitroanilide.

The enzymic properties of urokinase (EC 3.4.21.31) were studied. The kinetic parameters of hydrolysis of 5-oxo-Pro-Gly-Arg-NA were determined in the pH range 5-9, at 25 degrees C and 37 degrees C. The reaction is affected by only one ionizing group of urokinase with pK 7.15 (25 degrees C) and pK 6.82 (37 degrees C). The results indicate that 5-oxo-Pro-Gly-Arg-NA is a good model substrate for studies of the conversion of plasminogen to plasmin. The Km values of the urokinase-catalysed hydrolysis of plasminogen and 5-oxo-Pro-Gly-Arg-NA are of the same order of magnitude. Plasmin catalyses the hydrolysis of 5-oxo-Pro-Gly-Arg-NA, but the Km value is several hundred times that of urokinase. Urokinase is shown not to react with good plasmin substrates, such as Bz-Arg-OEt and D-Val-Leu-Lys-NA, but is linearly competitively inhibited by 6-amino-hexanoic acid and trans-4-aminomethylcyclohexane-1-carboxylic acid.

Enzymatic properties of alpha 2-macroglobulin-proteinase complexes. Apparent discrimination between covalently and noncovalently bound trypsin by reaction with soybean trypsin inhibitor.

The binding of trypsin to alpha 2-macroglobulin, the appearance of free beta-cysteinyl thiol groups of the formed complexes, the steady-state kinetics of their enzymic hydrolysis of carbobenzoxy-L-valyl-glycyl-L-arginyl-4-nitroanilide and finally their reactions with soybean trypsin inhibitor leading to the formation of ternary alpha 2-macroglobulin-trypsin-soybean trypsin inhibitor complexes were investigated. Each alpha 2-macroglobulin molecule binds two trypsin tightly; the dissociation constants were found to be unmeasureably small, but the extent of formation of 1:1 and 1:2 complexes at different molar ratios of alpha 2-macroglobulin to trypsin as determined from the appearance of thiol groups clearly indicated that binding of trypsin to alpha 2-macroglobulin shows negative cooperativity. Binding of the first trypsin makes the access of the second less easy. The kinetic results showed a decrease of the kc/Km value of hydrolysis of the tripeptide substrate by approx. 4-fold compared to that of free trypsin for each alpha 2-macroglobulin-bound trypsin. Here no differences were seen between the bound trypsins. The analysis of the reactions between the alpha 2-macroglobulin-trypsin complexes and soybean trypsin inhibitor shows that ternary complexes do form, although slowly, and that two processes occur, not only when 1:2 complexes but also when 1:1 complexes react with soybean trypsin inhibitor. Soybean trypsin inhibitor apparently discriminates between two distinct binding modes of trypsin to alpha 2-macroglobulin, the covalently and the noncovalently alpha 2-macroglobulin-bound trypsins.

Pregnancy zone protein, a proteinase binding alpha-macroglobulin. Stopped-flow kinetic studies of its interaction with chymotrypsin.

Human pregnancy zone protein (PZP) is a major pregnancy-associated plasma protein, strongly related to alpha 2-macroglobulin (alpha 2M). The proteinase binding reaction of PZP is investigated using chymotrypsin as a model enzyme. The time-course of the interaction is studied by measuring the change in intrinsic protein fluorescence of PZP-chymotrypsin reaction mixtures as a function of time after rapid mixing in a stopped-flow apparatus. Titrations show the changes of fluorescence at equilibrium to correspond with the formation of a chymotrypsin-PZP(tetramer) species. The kinetic results show the formation of the species to take place in an overall second-order process dependent on the concentrations of chymotrypsin and of PZP(dimers), k = 5 x 10(5) M-1 x s-1. Reactions of PZP-thiol groups do not give rise to fluorescence changes. The fluorescence changes most likely reflect the formation of an intermediate with intact thiol esters. Further analysis of the kinetic results suggests that the chymotrypsin-PZP(tetramer) intermediate is formed in two reaction steps: (1) initially native PZP(dimers) are cleaved at bait regions by enzyme molecules, and that is the rate determining reaction of the fluorescence changes; (2) association with another PZP(dimer) or PZP(dimer)-chymotrypsin complex in a very fast reaction that leads to the formation of 1:1 -chymotrypsin-PZP(tetramer) intermediate, probably with intact thiol esters. The interactions studied apparently are established early in the path of the reaction and the fluorescence changes probably reflect noncovalent enzyme-PZP contacts, which are not changed when covalent binding occurs. Further, fluorescence changes are seen only in reactions of PZP with enzymes, not with methylamine.

The action of factor Xa on peptide p-nitroanilide substrates: substrate selectivity and examination of hydrolysis with different reaction conditions.

Kinetic parameters for the action of bovine Factor Xa (EC 3.4.21.22) on 25 commercially available peptide p-nitroanilides have been determined. The selectivity constant, kc/Km, ranges from 1.5 X 10(1) to 2 X 10(6) M-1 X s-1 for the poorest and the best substates, respectively. The best substrates for Factor Xa were identified as those with arginine in the P1 position, and glycine in the P2 position. Quantitative distinction between lysine and arginine in the P1 position and other amino acids in the P2-P4 positions of the substrate is reported from the changes in the kinetic parameters for substrates differing in only a single amino acid in these positions. Effect of NaCl and CaCl2 concentrations and temperature on the action of Factor Xa on selected substrates have been assessed. Km values for Factor Xa hydrolysis of most substrates are greater than 100 microM. Solubility of the substrates consequently restricts measurements of reaction velocities to concentrations lower than desirable for optimally determining kc. Comparison of these kinetic parameters for Factor Xa with those of thrombin (Lottenberg, R., Hall, J.A., Blinder, M., Binder, E. and Jackson, C.M. (1983) Biochim. Biophys. Acta 742,539-557) for these same substrates indicates that the greater hydrolytic efficiency of thrombin is due primarily to lower Km values.