Simulation dataset of thermal and epithermal neutron self-shielding correction factors for 186W(n,γ)187W reaction rate experiments using tungsten foil targets.

In the experiments of neutron interaction with research samples, the incident neutron energy spectrum, distribution inside the irradiating sample volume, is affected by the unexpected neutron self-shielding effects. The nature of these effects is due to the formation and thickness of the irradiating sample, which significantly causes neutron self-absorption and multiple scattering inside the sample volume. The datasets presented in this article showed the thermal (Gth) and epithermal (Gepi) neutron self-shielding correction factors for the 186W(n,γ)187W neutron capture reaction rate in irradiating tungsten (W) foil samples with different thicknesses. The simulations were performed for three models of surface neutron source's geometries and relative orientations of the irradiating foil samples of isotropic cylinder surface neutron source with foil sample along to the center line, isotropic cylinder neutron source with foil sample flat to the center line, and isotropic spherical neutron source with foil sample placed at the center point. The range of sample thicknesses was from 10 µm to 2.5 mm. The uncertainties for each data point are also reported in the data table, making it more convenient for reuse in related experiments or evaluations.

Cryogenic spectrometer for measuring the far-IR to millimeter-wave absorptivity of cosmic analog dusts.

We report on the design, construction, and performance of a custom apparatus built to measure the frequency- and temperature-dependent absorptivity of millimeter-wave light by cosmic analog dusts. We highlight the unique challenges faced as well as a few key innovations that are part of the instrument. Among those is an ultra-compact Fourier transform spectrometer. We have measured its effective frequency range and FWHM resolution to be 150-2100 GHz and ∼45GHz, respectively. Another innovation is a cold sample positioner whose temperature can be controlled within the range of 3.7-50 K. The use of a pulse-tube cryocooler results in a pulse-synchronous signal that dominates the detector (bolometer) signal. Methods used to address that challenge are also presented.

The schizophrenia- and autism-associated gene, transcription factor 4 regulates the columnar distribution of layer 2/3 prefrontal pyramidal neurons in an activity-dependent manner.

Disruption of the laminar and columnar organization of the brain is implicated in several psychiatric disorders. Here, we show in utero gain-of-function of the psychiatric risk gene transcription factor 4 (TCF4) severely disrupts the columnar organization of medial prefrontal cortex (mPFC) in a transcription- and activity-dependent manner. This morphological phenotype was rescued by co-expression of TCF4 plus calmodulin in a calcium-dependent manner and by dampening neuronal excitability through co-expression of an inwardly rectifying potassium channel (Kir2.1). For we believe the first time, we show that N-methyl-d-aspartate (NMDA) receptor-dependent Ca2+ transients are instructive to minicolumn organization because Crispr/Cas9-mediated mutation of NMDA receptors rescued TCF4-dependent morphological phenotypes. Furthermore, we demonstrate that the transcriptional regulation by the psychiatric risk gene TCF4 enhances NMDA receptor-dependent early network oscillations. Our novel findings indicate that TCF4-dependent transcription directs the proper formation of prefrontal cortical minicolumns by regulating the expression of genes involved in early spontaneous neuronal activity, and thus our results provides insights into potential pathophysiological mechanisms of TCF4-associated psychiatric disorders.

hCG-dependent regulation of angiogenic factors in human granulosa lutein cells.

As prerequisite for development and maintenance of many diseases angiogenesis is of particular interest in medicine. Pathologic angiogenesis takes place in chronic arthritis, collagen diseases, arteriosclerosis, retinopathy associated with diabetes, and particularly in cancers. However, angiogenesis as a physiological process regularly occurs in the ovary. After ovulation the corpus luteum is formed by rapid vascularization of initially avascular granulosa lutein cell tissue. This process is regulated by gonadotropic hormones. In order to gain further insights in the regulatory mechanisms of angiogenesis in the ovary, we investigated these mechanisms in cell culture of human granulosa lutein cells. In particular, we determined the expression and production of several angiogenic factors including tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), Leptin, connective tissue growth factor (CTGF), meningioma-associated complimentary DNA (Mac25), basic fibroblast growth factor (bFGF), and Midkine. In addition, we showed that human chorionic gonadotropin (hCG) has distinct effects on their expression and production. hCG enhances the expression and production of TIMP-1, whereas it downregulates the expression of CTGF and Mac25. Furthermore it decreases the expression of Leptin. Our results provide evidence that hCG determines growth and development of the corpus luteum by mediating angiogenic pathways in human granulosa lutein cells. Hence we describe a further approach to understand the regulation of angiogenesis in the ovary.

Arabitol dehydrogenase as a selectable marker for rice.

Arabitol dehydrogenase has been adapted for use as a plant selectable marker. Arabitol is a five-carbon sugar alcohol that can be used by E. coli strain C, but not by the laboratory K12 strains. The enzyme converts the non-plant-metabolizable sugar arabitol into xylulose, which is metabolized by plant cells. Rice was transformed with a plant-expression-optimized synthetic gene using Biolistic-mediated transformation. Selection on 2.75% arabitol and 0.25% sucrose yielded a transformation efficiency (9.3%) equal to that obtained with hygromycin (9.2%). Molecular analyses showed that the atlD gene was integrated into the rice genome of selected plants and was inherited in a Mendelian manner. This study indicates that arabitol could serve as an effective means of plant selection.

Brassinolide improves embryogenic tissue initiation in conifers and rice.

Somatic embryogenesis (SE), the most promising technology for the large-scale production of high-value coniferous trees from advanced breeding and genetic engineering programs, is expected to play an important role in increasing productivity, sustainability, and the uniformity of future U.S. forests. To be successful for commercial use, SE technology must work with a variety of genetically diverse trees. Initiation in loblolly pine ( Pinus taeda L.), our main focus species, is often recalcitrant for desirable genotypes. Initiation percentages of loblolly pine, Douglas-fir [ Pseudotsuga menziesii (Mirb.) Franco], and Norway spruce ( Picea abies L., Karst.) were improved through the use of brassinolide. Brassinosteroids, which include brassinolide, are a relatively new group of natural plant growth regulators that are found in many plant species. They have been shown to have diverse, tissue-specific, and species-specific effects, including the stimulation of cell elongation and ethylene production and increasing resistance to abiotic stress. In our media, brassinolide was effective at concentrations ranging from 0.005-0.25 micro M. Using control medium (no brassinolide) and brassinolide-supplemented (0.1 micro M) medium, we achieved improved initiation percentages in loblolly pine, Douglas-fir, Norway spruce, and rice-15.0% to 30.1%, 16.1% to 36.3%, 34.6% to 47.4%, and 10%, respectively. Brassinolide increased the weight of loblolly pine embryogenic tissue by 66% and stimulated initiation in the more recalcitrant families of loblolly pine and Douglas-fir, thus compensating somewhat for genotypic differences in initiation. Initiation percentages in loblolly pine were improved through the combination of modified 1/2-P6 salts, 50 mg/l activated carbon (AC), adjusted levels of Cu and Zn (to compensate for adsorption by AC), 1.5% maltose, 2% myo-inositol (to raise the osmotic level, partially simulating the megagametophyte environment), 500 mg/l casamino acids, 450 mg/l glutamine, 2 mg/l alpha-naphthaleneacetic acid, 0.63 mg/l 6-benzylaminopurine, 0.61 mg/l kinetin, 3.4 mg/l silver nitrate, 10 micro M cGMP, 0.1 micro M brassinolide, and 2 g/l Gelrite. Across 12 open-pollinated families of loblolly pine, initiation percentages ranged from 2.5% to 50.7%, averaging 22.5%.

Rate of coronary vascularization during embryonic chicken development is influenced by the rate of myocardial growth.

OBJECTIVE: We tested the hypothesis that the degree of coronary microvessel formation in the embryonic heart is regulated by the magnitude of myocardial growth. METHODS: The outflow tract of Hamburger-Hamilton stage 21 chicken hearts (prior to the onset of coronary vasculogenesis) was constricted in ovo with a loop of 10-0-nylon suture, and the hearts were studied at stages 29 and 36. RESULTS: At stage 29 ventricular mass was 64% greater in the pressure-overloaded than in the hearts of sham-operated controls, but vascular volume density and numerical density, determined by electron microscopic morphometry, were identical. As demonstrated by histological morphometric evaluation, the compact region of the left ventricle at stage 29 was 43% thicker than the shams. However, by stage 36 heart mass, thickness of the compact region, and overall wall thickness (demonstrated by scanning electron microscopy) were significantly less than in the sham group of this stage, but vascular volume density was virtually identical in the two groups. Formation of the two main coronary arteries was clearly impeded in the banded hearts, i.e., the coronaries were stunted in their development or failed to completely form coronary ostia. CONCLUSIONS: Vascular growth is proportional to myocardial growth in the embryonic, overloaded heart, but the persistence of the pressure overload results in a failure of or severe limitations in coronary artery development. These data support the hypothesis that vascular growth during this period of development is regulated, at least in part, by the rate and magnitude of myocardial growth.

Locus coeruleus neurons: cessation of activity during cataplexy.

Cataplexy, a symptom of narcolepsy, is a loss of muscle tone usually triggered by sudden, emotionally significant stimuli. We now report that locus coeruleus neurons cease discharge throughout cataplexy periods in canine narcoleptics. Locus coeruleus discharge rates during cataplexy were as low as or lower than those seen during rapid-eye-movement sleep. Prazosin, an alpha1 antagonist, and physostigmine, a cholinesterase inhibitor, both of which precipitate cataplexy, decreased locus coeruleus discharge rate. Our results are consistent with the hypothesis that locus coeruleus activity contributes to the maintenance of muscle tone in waking, and that reduction in locus coeruleus discharge plays a role in the loss of muscle tone in cataplexy and rapid-eye-movement sleep. Our results also show that the complete cessation of locus coeruleus activity is not sufficient to trigger rapid-eye-movement sleep in narcoleptics.

Doppler echocardiography as a predictor of pregnancy outcome in the presence of aortic stenosis: A case report.

Aortic stenosis in pregnancy can be a life-threatening condition, but fortunately it is rare. In the modern era, careful obstetric and cardiologic monitoring, particularly through echocardiography, have improved fetal and maternal outcomes. However, a test that could predict outcome has not been available for patients with aortic stenosis who seek prepregnancy counseling. We report a case in which exercise Doppler echocardiography was used to predict cardiac function and maximal gradients in a woman with a bicuspid aortic valve who wished to become pregnant.

Discovery of a novel series of potent and selective substrate-based inhibitors of p60c-src protein tyrosine kinase: conformational and topographical constraints in peptide design.

On the basis of the efficient substrate for p60c-src protein tyrosine kinase (PTK) YIYGSFK-NH2 (1) (Km = 55 microM) obtained by combinatorial methods, we have designed and synthesized a series of conformationally and topographically constrained substrate-based peptide inhibitors of this enzyme, which showed IC50 values in the low-micromolar range (1-3 microM). A "rotamer scan" was performed by introducing the four stereoisomers of beta-Me(2')Nal in the postulated interaction site of the peptide inhibitor 23(IC50 = 1.6 microM). This substitution led to selective and potent inhibitors of p60c-src PTK; however, no substantial difference in potency was observed among them. This and the results of the "stereochemical scan" performed at residues 2 and 7 of 3 (peptides 19-21), which form the disulfide bond, may suggest that the enzyme active site does not have rigid topographic requirements and thus is able to achieve important conformational changes to bind the ligand as long as the pharmacophore pattern in the inhibitor is conserved. Two new potent iodo-containing nonphosphorylatable tyrosine analogues were also incorporated into our lead inhibitory sequence 23, producing the most potent inhibitors for p60c-src PTK identified thus far in our studies. Compounds 29 and 30 exhibit IC50 values of 0.13 and 0.54 microM, respectively. Peptide 29 is 420-fold more potent than the parent peptide 1. Selectivity studies of peptides 23-30 toward p60c-src, Lyn, and Lck PTK showed in general high Lyn/Src and moderate Lck/Src selectivity ratios. We found that the chi1 space constraints of the specialized amino acids, introduced at position 3 of the peptide lead 23, were not as important as the configuration of the Calpha of that residue to recognize the subtle chemical environment surrounding the active site of Src and Lck PTK, as reflected on the obtained Lck/Src selectivity ratios.

Effect of complexes of ADP and phosphate analogs on the conformation of the Cys707-Cys697 region of myosin subfragment 1.

Recent crystallographic studies have suggested structural differences between the complexes of S1.Mg.ADP with the phosphate analogs aluminium fluoride (AlF4-), vanadate (VO(4)3-) and beryllium fluoride (BeFx) [Fisher, A. J., Smith, C. A., Thoden, J. B., Smith, R., Sutoh, K., Holden, H. M. & Rayment, I. (1995) Biochemistry 34, 8960-8972; Smith, R. & Rayment, I. (1996) Biochemistry 35, 5404-5417]. In this work, chemical modifications, namely labeling of Cys707 (the reactive SH1 thiol) and Cys707-Cys697 (SH1-SH2) cross-linking, were used to compare the S1.ADP.BeFx, S1.ADP. AlF4- and S1.ADP-VO(4)3- complexes with specific states of the myosin-ATPase pathway. Modification of Cys707 with the fluorescent monofunctional reagents 7-diethylamino-3-(4'-maleimidylphenyl)-4-methylcoumarin and N-iodoacetyl-N'-(5-sulfo-1-naphtyl)ethylenediamine has shown that the reactivity of the SH1 group depends on the nucleotide bound to S1. The observed rates of Cys707 modification at 20 degrees C lead to the conclusion that S1.ADP.BeFx is similar to S1*.ATP, while S1.ADP.AlF4- and S1.ADP.VO(4)3- are more similar to S1**.ADP.Pi. The conformations of the analog states were also compared by monitoring the dissociation of the fluorescent nucleotide analog 1-N6-ethenoadenosine diphosphate (ADP[C2H2]) from the active site of Cys707-modified (by N-ethylmaleimide) and Cys707-Cys697-cross-linked (by N,N'-p-phenylene dimaleimide) S1.ADP[C2H2].AlF4- and S1.ADP[C2H2]. BeFx. Our results suggest that the conformations of the S1.ADP.AlF4-, S1.ADP.VO(4)3- and S1.ADP.BeFx complexes in the Cys707-Cys697 region are distinct from each other, with the former two at least partially resembling the S1**.ADP.Pi state, while the latter is similar to the prehydrolyzed S1*.ATP state.

Interactions of the subunits of smooth muscle myosin phosphatase.

Myosin phosphatase from smooth muscle consists of a catalytic subunit (PP1c) and two non-catalytic subunits, M130 and M20. Interactions among PP1c, M20, and various mutants of M130 were investigated. Using the yeast two-hybrid system, PP1c was shown to bind to the NH2-terminal sequence of M130, 1-511. Other interactions were detected, i.e. PP1c to PP1c, M20 to the COOH-terminal fragment of M130, and dimerization of the COOH-terminal fragment of M130. Mutants of M130 were constructed to localize the PP1c and light chain binding regions. Results from the two-hybrid system indicated two binding sites for PP1c on M130: one site in the NH2-terminal 38 residues and a weaker site(s) in the ankyrin repeats region. Inhibition of PP1c activity with phosphorylase a by the M130 mutants also was consistent with the assignment of these two sites. Overlay assays showed binding of phosphorylated light chain to the ankyrin repeats, probably in the COOH-terminal repeats. Activation of PP1c with phosphorylated light chain required binding sites for PP1c and substrate, plus an additional sequence COOH-terminal to the ankyrin repeats. Thus, activation of phosphatase and binding of PP1c and substrate are properties of the NH2-terminal one-third of M130.

Complexes of myosin subfragment-1 with adenosine diphosphate and phosphate analogs: probes of active site and protein conformation.

Previous work has revealed phosphate-dependent differences in the complexes formed from myosin subfragment-1 with adenosine diphosphate (S1.ADP) and aluminum fluoride (AlF4-) or beryllium fluoride (BeFx) [Phan and Reisler, Biophys. J., 66 (1994) A78], with the former resembling more the S1**.ADP.Pi state while the latter resembles more the S1.ATP state. In this work, the conformations of the S1.epsilon ADP.AlF4- and S1.epsilon ADP.BeFx, complexes were examined by nucleotide chase and collisional quenching experiments. epsilon ADP release from S1.epsilon ADP.AlF4- was slower than that from S1.epsilon ADP.BeFx. However, acrylamide titrations of S1.epsilon ADP.AlF4- and S1.epsilon ADP.BeFx showed little difference in nucleotide protection from quenching between the two complexes. This contrasts with the earlier observation on phosphate analog-dependent changes in the reactivity of the SH1 group on S1. To confirm phosphate-related perturbation of the SH1-SH2 sequence, emission spectra of fluorescein (IAF)-labeled SH1 and IANBD-labeled SH2 were recorded for S1 complexes with nucleotides and phosphate analogs. Considerable differences were found between the BeFx and AlF4- complexes with S1.MgADP for both SH1- and SH2-labeled proteins. These results are consistent with a recent crystallographic study of S1 complexes with ADP and phosphate analogs [Fisher et al., Biophys. J., 68 (1995) 19S] and the idea that the opening of the nucleotide cleft on S1 does not change much during ATP hydrolysis [Franks-Skiba et al., Biochemistry, 33 (1994) 12720], while significant changes in the SH1-SH2 region accompany phosphate cleavage.

Successful thrombolysis for prosthetic pulmonary valve obstruction.

Thrombosis is a serious complication of prosthetic heart valve operations. In recent years, systemic thrombolysis has emerged as a suitable alternative to surgery. Experience with thrombosis of pulmonary prosthetic valves is very limited. We report a case of successful administration of intravenous streptokinase for thrombosis of a St. Jude Medical prosthetic valve 3 weeks after pulmonary valve replacement.

Extensively methylated myosin subfragment-1: examination of local structure, interactions with nucleotides and actin, and ligand-induced conformational changes.

The atomic structure of myosin subfragment-1 (S1) has been recently solved for crystals of extensively methylated S1 [Rayment et al. (1993) Science 261, 50-58]. In this study, the effect of such a modification on S1 structure and function was examined. According to the far- and near-ultraviolet CD spectra, the methylation does not affect the secondary structure of S1 but causes limited changes in its tertiary structure. The methylation significantly decreases the affinity of S1 for actin in rigor and, to a lesser degree, that of S1 to actin in the presence of MgATP gamma S. This modification, like the trinitrophenylation of Lys-83, accelerates the dissociation of a nucleotide trapped on S1 either by phosphate analogs or by cross-linking of the SH1 and SH2 thiols. Methylation strongly impairs the coupling between the actin- and nucleotide-binding sites as revealed by the reduced effect of actin on the release of epsilon ADP from the active site. It also causes a complete loss of in vitro motility of actin filaments over methylated HMM. In addition to this, methylation also impairs the communication between other sites on S1 including that between the nucleotide-binding site and SH1, and the actin-binding site and the 27/50 kDa junction and a site at 74 kDa from the N-terminus of S1. These changes are revealed in SH1 modification, thermolysin digestion, and vanadate-dependent photocleavage experiments, respectively. The increased rate of thermal denaturation of S1 and the loss of S1 protection by ADP and actin from this process also indicate flawed communications in methylated S1. It is concluded that these relatively mild but numerous and important changes impair the function of methylated S1.

Dynamic properties of actin. Structural changes induced by beryllium fluoride.

Beryllium fluoride (BeFx) has been widely used as a phosphate analogue in nucleotide-binding proteins. It was found to bind tightly to F- but not G-actin (Combeau C., and Carlier M. F. (1988) J. Biol. Chem. 263, 17429-17436) and to affect the three-dimensional structure of filaments by stabilizing the subdomain 2 region of the actin promoter (Orlova, A., and Egelman, E. H. (1992) J. Mol. Biol. 227, 1043-1053). In this work we examined the BeFx-induced structural and functional changes in G- and F-actin by using proteolysis, chemical modifications, ATPase, and in vitro motility assays. The results of proteolysis studies show that BeFx binds also to MgADP-G-actin and renders its subdomain 2 region more similar to that in MgATP-G-actin. This is manifested in enhanced subtilisin and decreased tryptic digestions in subdomain 2 of G-actin. BeFx had a strong effect on the proteolysis of MgADP-F-actin: both the tryptic and subtilisin digestions in subdomain 2 were completely inhibited. Significant protection against proteolysis in this region was observed even at 1:14 molar ratios of BeFx to actin indicating cooperative effects on the structure of the actin filament. A similar although milder effect of phosphate on the proteolysis of F-actin suggests that BeFx acts as a phosphate analogue in this system. BeFx also induces changes in the subdomain 1 region of F-actin. This is revealed via reduced rates of Cys-374 alkylation with 7-diethylamino-3-(4'-maleimidylphenyl)-4-methylcoumarin and an increased subtilisin cleavage near the C terminus of actin in the presence of BeFx. The BeFx-induced structural changes in actin have little effect on its interactions with myosin. BeFx inhibits only slightly the actin-activated ATPase activity of S1 by decreasing Vmax without affecting KM. Additionally, the binding of BeFx to actin does not change the sliding velocity of actin filaments in the in vitro motility assays. The BeFx-induced specific and distinct changes in G- and F-actin point to the dynamic nature of actin structure and the local differences between monomeric and polymeric forms of actin.

Aluminum fluoride interactions with troponin C.

The increasing interest in the metal ion aluminum fluoride and beryllium fluoride complexes as phosphate analogs in the myosin ATPase reaction and in muscle fiber studies prompted the examination of their interactions with the regulatory system of troponin and tropomyosin. In this work, the effects of these metal ion analogs on the spectral properties of the Ca(2+)-binding subunit of troponin, troponin C (TnC), were examined. In contrast to beryllium fluoride which did not change the spectral properties of TnC, aluminum fluoride binding induced an increase in both the alpha-helicity and the tyrosine fluorescence of TnC and exposed a hydrophobic region on this protein for fluorescent probe binding. Aluminum fluoride also reduced the Ca2+ and/or Mg(2+)-induced changes on TnC. These results indicate a direct interaction of aluminum fluoride with TnC and merit consideration in designing muscle fiber experiments with this phosphate analog.

Kinetic and equilibrium analysis of the interactions of actomyosin subfragment-1.ADP with beryllium fluoride.

The hypothesis that the stable ternary complex formed between myosin subfragment-1, MgADP and beryllium fluoride (BeF3-), denoted S-1 not equal to .ADP.BeF3-, is an analog of the intermediate state S-1**.ADP.P(i) has been tested in this work by examining the interactions of S-1 not equal to .ADP.BeF3- with actin. Equilibrium binding measurements revealed that actin bound weakly to the S-1 not equal to .ADP.BeF3- complex (Ka = 10(4) M-1) in the presence of 40 mM KCl. The stability of this complex was strongly salt-dependent. The association constant of BeF3- to the acto-S-1.ADP complex (KBe approximately 10(3) M-1) was 100-fold weaker than its binding to the S-1.ADP complex. While inhibiting the S-1 ATPase strongly, BeF3- had no effect on the Vmax value (10 +/- 1.0 s-1) of the actin-activated ATPase of S-1. The rates of BeF3- binding and dissociation from the acto-S-1.ADP.BeF3- complex were determined by stopped-flow measurements. The hyperbolic dependence of the rates of BeF3- binding to acto-S-1.ADP (kobs) on BeF3- concentrations suggested that the acto-S-1.ADP.BeF3- complex was formed in at least two steps: binding followed by isomerization. The binding constant was 1.2 x 10(3) M-1, and the maximum kobs was 2.5 s-1. The dissociation of BeF3- from the acto-S-1.ADP.BeF3- complex was monitored via decrease in the fluorescence of 1-N6-ethenoadenosine diphosphate (epsilon ADP). The fluorescence decrease fitted two exponential terms.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of myosin ATPase by beryllium fluoride.

Inhibition of the myosin subfragment 1 (S-1) ATPase activity by beryllium fluoride was studied directly in the presence of MgATP and following preincubation of samples with MgADP. In both cases, the rates of inhibition were very slow, with kapp = 0.5 and 58 M-1 s-1, respectively, in analogy to the rates of inhibition of myosin ATPase by vanadate [Goodno, C. C. (1979) Proc. Natl. Acad. Sci. U.S.A. 76, 2620-2624]. The very different rates of inhibition in the presence of MgATP and on preincubation with MgADP suggested that beryllium fluoride binds to the M.ADP state of myosin. The slow inhibition rates and the nonlinear dependence of the observed rates on beryllium fluoride concentration were consistent with a two-step inhibition process involving a rapid binding equilibrium to yield a collisional complex, M.ADP.BeF3-, and its slow isomerization into M++.ADP.BeF3-. A third, much slower, step was required to account for the conversion of the stable M++.ADP.BeF3- to a virtually irreversibly inhibited complex. Kinetic description of the inhibition pathway was derived from the observed rates of inhibition of myosin ATPase, information on the binding of beryllium fluoride to M.ADP, and measurements of epsilon ADP chase from M++.epsilon ADP.BeF3-. The isomerization rate and equilibrium constants were 1.4 x 10(-2) s-1 and 50, respectively, and the overall binding constant of beryllium fluoride to M.ADP was 5 x 10(5) M-1. The inhibitory complex showed a 16% enhancement to tryptophan fluorescence of S-1 and a reduced quenching of epsilon ADP by acrylamide. It is concluded that M++.ADP.BeF3- is analogous to the M++.ADP.Vi and M**.ADP.Pi states of myosin.