Medical research in Israel and the Israel biomedical database.

The data collected for the second edition of the Directory of Medical Research in Israel and the Israel Biomedical Database have yielded very relevant information concerning the distribution of investigators, publication activities and funding sources. The aggregate data confirm the findings of the first edition published in 1996 [2]. Those facts endorse the highly concentrated and extensive nature of medical research in the Jerusalem area, which is conducted at the Hebrew University and its affiliated hospitals. In contrast, Tel Aviv University, whose basic research staff is about two-thirds the size of the Hebrew University staff, has a more diffuse relationship with its clinical staff who are located at more than half a dozen hospitals. Ben-Gurion University in Beer Sheva and the Technion in Haifa are smaller in size, but have closer geographic contact between their clinical and basic research staff. Nonetheless, all the medical schools and affiliated hospitals have good publication and funding records. It is important to note that while some aspects of the performance at basic research institutions seem to be somewhat better than at hospitals, the records are actually quite similar despite the greater burden of clinical services at the hospitals as compared to teaching responsibilities in the basic sciences. The survey also indicates the substantial number of young investigators in the latest survey who did not appear in the first survey. While this is certainly encouraging, it is also disturbing that the funding sources are apparently decreasing at a time when young investigators are attempting to become established and the increasing burden of health care costs precludes financial assistance from hospital sources. The intensity and undoubtedly the quality of medical research in Israel remains at a level consistent with many of the more advanced western countries. This conclusion is somewhat mitigated by the fact that there is a decrease in available funding and a measurable decrease in scholarly activity at a time when a new, younger generation of investigators is just beginning to become productive. In closing, we wish to stress that the collection of data for the Biomedical Database is a continuing project and we encourage all medical researches who may not have contributed relevant information to write to the Office of the Chief Scientist or contact the office by email.

Emerging infectious diseases: a cause for concern.

As the twenty-first century begins it becomes increasingly apparent that the twentieth century, which opened with the promise of the eradication of most infectious diseases, closed with the specter of the reemergence of many deadly infectious diseases that have a rapidly increasing incidence and geographic range. Equally if not more alarming is the appearance of new infectious diseases that have become major sources of morbidity and mortality. Among recent examples are HIV/AIDS, hantavirus pulmonary syndrome, Lyme disease, hemolytic uremic syndrome (caused by a strain of Escherichia coli), Rift Valley fever, Dengue hemorrhagic fever, malaria, cryptosporidiosis, and schistosomiasis. The reasons for this situation are easily identified in some cases as associated with treatment modalities (permissive use of antibiotics), the industrial use of antibiotics, demographic changes, societal behavior patterns, changes in ecology, global warming, the inability to deliver minimal health care and the neglect of well-established public health priorities. In addition is the emergence of diseases of another type. We have begun to characterize the potential microbial etiology of what has historically been referred to as chronic diseases.

Research in medical sciences in Israel, 1994-97, an overview of institutions, investigators, and funding.

This is a summary of the second effort to characterize medical research activities in Israel (1994-97). The study relies on responses to a questionnaire sent to all participants in the first survey (1991-94) and to additional, including younger, investigators. There were 1450 direct responses this time versus 1088 the first time. The distribution of investigators is 60% hospital-based versus 40% in basic sciences. The average number of funding sources per investigator has decreased from 2.6 to 2.2; however the percentage of funded investigators has increased from 60% to 70%. The greatest concentration of funded investigators remains at the Hebrew University and Hadassah Medical Center, followed by Tel Aviv University and associated hospitals. The Chief Scientist's Office of the Ministry of Health funds more than 40% of funded investigators (more than double the next highest funding source, the US-Israel Binational Fund), followed in decreasing order by the National Academy of Sciences, Ministry of Science, German-Israeli Foundation for Scientific Research and Development (GIF), Israel Cancer Society, and NIH. The percentage of funded hospital-based investigators has increased from 59% to 62%; however, funding remains higher for basic science investigators. There has been an increase in the percentage of Ph.D.'s working in a hospital environment and an interesting anomaly is that the nonacademic-affiliated Ph.D. investigators in hospitals have been successful in receiving funding, so now a greater percentage of nonacademic hospital staff are funded than academic staff (69% versus 61%).

Comparative thermodynamic elucidation of the structural stability of thermophilic proteins.

Differential scanning calorimetry, circular dichroism, and visible absorption spectrophotometry were employed to elucidate the structural stability of thermophilic phycocyanin derived from Cyanidium caldarium, a eucaryotic organism which contains a nucleus, grown in acidic conditions (pH 3.4) at 54 degrees C. The obtained results were compared with those previously reported for thermophilic phycocyanin derived from Synechococcus lividus, a procaryote containing no organized nucleus, grown in alkaline conditions (pH 8.5) at 52 degrees C. The temperature of thermal unfolding (t(d)) was found to be comparable between C. caldarium (73 degrees C) and S. lividus (74 degrees C) phycocyanins. The apparent free energy of unfolding (DeltaG([urea]=0)) at zero denaturant (urea) concentration was also comparable: 9.1 and 8.7 kcal/mole for unfolding the chromophore part of the protein, and 5.0 and 4.3 kcal/mole for unfolding the apoprotein part of the protein, respectively. These values of t(d) and DeltaG([urea]=0) were significantly higher than those previously reported for mesophilic Phormidium luridum phycocyanin (grown at 25 degrees C). These findings revealed that relatively higher values of t(d) and DeltaG([urea]=0) were characteristics of thermophilic proteins. In contrast, the enthalpies of completed unfolding (DeltaH(d)) and the half-completed unfolding (DeltaH(d)) 1 2 for C. caldarium phycocyanin were much lower than those for S. lividus protein (89 versus 180 kcal/mole and 62 versus 115 kcal/mole, respectively). Factors contributing to a lower DeltaH(d) in C. caldarium protein and the role of charged groups in enhancing the stability of thermophilic proteins were discussed.

Newborn seroprevalence study: methods and results.

For the 28-month period, November 30, 1987 through March 31, 1990, 653,117 blood specimens obtained on all newborn infants in New York State for detection of metabolic disorders were also analyzed for HIV serologic status. The overall seroprevalence rate was 0.66 percent: 1.24 percent in New York City and 0.17 percent in New York State exclusive of New York City. Rates of seropositivity were highest in the Bronx (1.72 percent) and Manhattan (1.59 percent). Outside of New York City, HIV seropositivity was concentrated in certain areas. Sixty-four zip codes with two or more seropositives and an HIV seroprevalence rate twice the average outside of New York City contained 65 percent of the HIV seropositives but only 16 percent of the newborns tested. Newborn seropositivity increased with maternal age. In New York City, the seroprevalence rates increased from 0.16 percent (1 in 624) for 14-year-olds to 1.41 percent (1 in 71) for 24-year-olds, a ninefold rise. This survey has provided the impetus for a number of preventive initiatives.

A methodology for determination of phospholipids.

A simple method for the determination of phospholipids in an aqueous dispersion and in amniotic fluid was developed. The procedure is based on the observation that dispersed phospholipids promoted the solubilization of an insoluble dye--detergent complex. The solubilization of the complex between the negatively charged dye, Coomassie brilliant blue (CBB), and a positively charged detergent, cetyltrimethylammonium bromide (CTAB), produced a blue solution having a visible absorbance maximum above 600 nm. A linear increase in absorbance intensity occurs with an increase in phospholipid concentration. An assay using the CBB-CTAB reagent adsorbed on 3-mm glass beads is used to estimate total dispersed phospholipids between 2 and 25 micrograms/ml. Thereby, a two-phase water-methanol-chloroform system is formed. The products of zwitterionic phospholipids (such as phosphatidylcholine and phosphatidylethanolamine) partition to the organic phase while the dye complex solubilized in anionic phospholipids (such as phosphatidylglycerol and phosphatidylinositol) partitions to the aqueous phase. This procedure results in a convenient, sensitive, and rapid method for the simultaneous determination of the total phospholipid, zwitterionic phospholipid, and anionic phospholipid concentrations. Application of the new assay for determination of phospholipids in amniotic fluid is described.

The Chromophore and Polypeptide Composition of Aplysia Ink.

The composition of the ink of the sea hare, Aplysia, was studied in regard to its tetrapyrrole and polypeptide content. The ink was separated into three pigment components by both thin-layer and gel filtration chromatography. These three pigments have distinctive visible absorption spectra, and--by comparison with known tetrapyrroles--we have demonstrated that they are derived from the three tetrapyrrole chromophores (bilins) found on the biliproteins of certain red algae, which constitute a portion of the Aplysia diet. The red component is phycourobilin; the purple is phycoerythrobilin; and the blue is phycocyanobilin. Sodium dodecyl sulfate gel electrophoresis experiments were also performed. The results of these experiments showed several polypeptides, and major bands at 78,000 and 61,000 molecular weight were noted. Biliproteins, at most, would be minor components of the ink.

Monitoring human immunodeficiency virus type 1-infected patients by ratio of antibodies to gp41 and p24.

Antibody responses of 85 patients to human immunodeficiency virus type 1 antigens were quantitated by densitometric analysis of Western blot (immunoblot) assays. All patients had been classified into the following three clinical categories: asymptomatic (ASY), acquired immunodeficiency syndrome (AIDS)-related complex (ARC), or AIDS. Fifty of the patients were monitored for 6 to 29 months. The gp41/p24 antibody ratio was examined in three studies. In the first study, initial specimens from each patient were analyzed. The mean gp41/p24 antibody ratios were 1.5 (ASY), 3.2 (ARC), and 5.4 (AIDS). Of ASY patients, 79% had antibody ratios of less than 2.0. In contrast, 72% of patients with AIDS had ratios of greater than or equal to 2.0. In the second study, serially obtained specimens from ASY, ARC, and AIDS patients were analyzed. These patients were further grouped according to progression of their clinical condition. Of ASY patients whose clinical condition progressed to ARC, 80% consistently had ratios of greater than or equal to 2.0. Of ARC patients whose clinical condition progressed to AIDS, 71% consistently had ratios of greater than or equal to 2.0. Of AIDS patients who died during the study, 100% consistently had ratios of greater than or equal to 2.0. No patients were treated with azidothymidine during the first two studies. In the third study, AIDS patients were monitored before and during treatment with azidothymidine. During treatment, ratios stabilized or improved transiently in five of seven patients. In these three studies, a gp41/p24 antibody ratio of less than 2.0 correlated with a benign clinical state and a ratio of greater than or equal to 2.0 correlated with AIDS or progression to AIDS.

Interaction of C-phycocyanin with lipid monolayers under nitrogen and in the presence of air.

The surface interaction of C-phycocyanin with lipids was studied using the monolayer technique. The surface activity of the protein was found to be higher at the lipid-water interface than at the nitrogen-water interface, particularly at high surface pressures of the lipid monolayer. The maximum initial surface pressures beyond which phycocyanin could not penetrate the dipalmitoylphosphatidylcholine and monogalactosyldiglycerol monolayers were 27 and 30 mN m-1, respectively. Below these values the protein demonstrated preferential interaction with the monogalactosyldiglycerol monolayer. The surface properties of the unfolded protein at pH 2.5 at the lipid-water interface were compared with those of the protein at pH 7.0. Higher affinity of the three-dimensional structure of the protein to lipid monolayers was observed, in particular by high subphase protein concentration. When the lipid films were subjected to oxidation stress by exposure to air, the surface properties of C-phycocyanin and dipalmitoylphosphatidylcholine were not greatly affected but the surface activity of monogalactosyldiacylglycerol was reduced dramatically by autoxidation. The oxidation of monogalactosyldiacylglycerol could not be prevented by the introduction of C-phycocyanin molecules at the lipid-water interface.

Densitometric analysis of Western blot (immunoblot) assays for human immunodeficiency virus antibodies and correlation with clinical status.

Western blot assays for antibodies directed against components of human immunodeficiency virus (HIV) associated with acquired immunodeficiency syndrome (AIDS) were examined with a densitometer and integrator. Antibody responses to seven HIV proteins were determined from the areas under the peaks of bands on blots from 430 seropositive individuals. Antibody responses corresponded qualitatively and quantitatively with clinical status. The Western blot assays examined were done on single specimens from individuals in one of four clinical states: asymptomatic with no risk factor identified, asymptomatic with risk factor(s) identified, AIDS-related complex, and AIDS. The ratios of gp41 antibody to p24 antibody and of gp41 antibody to total HIV antibodies increased, and the number of total HIV antibodies decreased progressively in these populations. Parameters were assigned to characterize the typical response found in AIDS: gp41 antibody/p24 antibody ratio, greater than or equal to 2.0; gp41 antibody/total HIV antibodies ratio, greater than or equal to 0.30; and number of total HIV antibodies, less than or equal to 25.0 signal units. Parameter match increased with progression of clinical status. These parameters were applied in a brief follow-up study of 34 HIV-infected asymptomatic individuals who developed AIDS-related complex or AIDS. Initial specimens showed a stronger correlation than our population data base had predicted, suggesting that the parameters have prognostic value. Densitometric analysis of antibody responses on Western blot assays of single or serial specimens should prove useful to physicians in staging and monitoring HIV-infected individuals and in predicting which individuals will progress to AIDS.

Kinetics of allophycocyanin's trimer-monomer equilibrium.

Kinetic studies of the dissociation of allophycocyanin trimers to monomers have been performed by using stopped-flow techniques. The dissociation was monitored by two techniques: by light scattering to observe the molecular weight changes directly and by 650-nm absorbance to observe the linkage of quaternary structure to spectra. The light-scattering experiments showed a simple exponential decay of trimers to monomers with a dissociation constant of 0.23 s-1. The absorption changes were complex, with two processes occurring. The faster absorption change appeared to be almost simultaneous with the molecular weight change (about 0.27 s-1) and was perhaps totally coordinated with it. The slower absorption change (0.071 s-1) was possibly a result of a conformational change in the chromophore arising during the conversion from newly dissociated monomers to equilibrium monomers.

Picosecond time-resolved fluorescence from detergent-free photosystem I particles.

Picosecond time-resolved fluorescence measurements have been taken on a detergent-free P700-enriched complex at room temperature isolated from the blue-green alga Phormidium luridum with a chlorophyll a to reaction center ratio of 100. Emission at greater than 665 nm is characterized by two exponential-decay components. A fast component, which dominates the initial decay with an average lifetime of 16 ps and 87% amplitude, is attributed to excitations in the core antenna chlorophyll-proteins, which are rapidly trapped by the primary electron donor, P700. A second component, with an average lifetime of 106 ps and 13% amplitude, is attributed to the peripheral antenna proteins. For 532-nm, 30-ps pulse excitation the results are virtually independent of fluence in the range of 2 x 10(12) to 4 x 10(16) photons/cm(2) and the oxidation state of P700. Addition of sodium dodecyl sulfate to 0.1% causes the second component's lifetime to increase by an average of a factor of 2.5. Only minor changes are observed in the first component's lifetime and the relative amplitudes of the two components. Two fractions isolated from the detergent-treated samples have also been examined. Our results indicate that excitation energy transfer within photosystem I is very efficient and that the excitation kinetics of the antennae may be limited by the trapping rate of P700 or strongly affected by the heterogeneity of the antennae.

Picosecond fluorescence of cryptomonad biliproteins. Effects of excitation intensity and the fluorescence decay times of phycocyanin 612, phycocyanin 645, and phycoerythrin 545.

The fluorescence of purified biliproteins (phycocyanin 645, phycocyanin 612, and phycoerythrin 545) from three cryptomonads, Chroomonas species, Hemiselmis virescens, and Rhodomonas lens, and C-phycocyanin from Anacystis nidulans has been time resolved in the picosecond region with a streak camera system having less than or equal to 2-ps jitter. The fluorescence lifetimes of phycocyanins from Chroomonas species and Hemiselmis virescens are 1.5 +/- 0.2 ns and 2.3 +/- 0.2 ns, respectively, regardless of the fluence of the 30 ps, 532-nm excitation pulse. (Fluence [or photons/cm2] = f intensity [photons/cm2s]dt.). In contrast, that of C-phycocyanin is 2.3 +/- 0.2 ns when the excitation fluence is 8.2 X 10(11) photons/cm2 and decreases to a decay approximated by an exponential decay time of 0.65 +/- 0.1 ns at 7.2 X 10(16) photons/cm2. The cryptomonad phycoerythrin fluorescence decay lifetime is also dependent on intensity, having a decay time of 1.5 +/- 0.1 ns at low fluences and becoming clearly biphasic at higher fluences (greater than 10(15) photons/cm2). We interpret the shortening of decay times for C-phycocyanin and phycoerythrin 545 in terms of exciton annihilation, and have discussed the applicability of exciton annihilation theories to the high fluence effects.

Effect of acetic acid on phycocyanin-phycoerythrocyanin mixture extracted from Anabaena variabilis.

Exposure of a phycocyanin-phycoerythrocyanin mixture extracted from Anabaena variabilis to sodium acetate, pH 3.8, ionic strength of 0.1, results in dissociation of the phycoerythrocyanin's beta subunit from its alpha subunit. The alpha subunit obtained by this method has a strong absorption transition at 508 nm. This transition is a consequence of the subunit's specific conformation, rather than of a new chromophore. The behavior of the phycocyanin-phycoerythrocyanin mixture in acetate buffers of variable compositions suggests that interactions which involve carboxylic amino acid residues play an important role, along with hydrophobic associations, in the association of phycoerythrocyanin subunits into monomers (alpha beta) and between this protein and phycocyanin. This work also indicates that the linkage between alpha and beta subunits of phycoerythrocyanin is labile and may be weaker than the association of these subunits with phycocyanin under acidic conditions.

Partial characterization of six chlorophyll a-protein complexes isolated from a blue-green alga by a nondetergent method.

The chlorophyll a-protein complexes of a blue-green alga, Phormidium luridum, are resolved in the absence of detergent, by the combination of a Sepharose 4B column and sucrose density gradient centrifugation, into six chlorophyll a-containing zones. These six complexes are termed here as F15, F25, F35, F40, F60, and F65, according to their appearance in the sucrose density gradient after centrifugation. The absorption spectra of these six isolated complexes are reported, as well as the fluorescence emission spectra at room temperature and liquid-nitrogen temperature. The F60 complex was enriched in Photosystem I, while the F35 and F40 complexes contained both Photosystem I and II. The F60 complex is the predominant band and accounts for about 49% of the total chlorophyll a of the cells. The extinction coefficient of this complex is determined to be 68.7 mM-1 cm-1 at 680 nm in 50 mM tris(hydroxymethyl)amino methane buffer at pH 8.0. In addition, the effect of the detergent, sodium dodecyl sulfate, on these spectra are also reported for comparison. The chemically induced difference spectra of F35, F40, and F60 complexes also indicate the presence of the reaction center, P700, of Photosystem I. These three complexes have been shown to contain P700 in a ratio of approximately one reaction center molecule per 100 light-harvesting chlorophyll molecules. The simple exposure of the F60 fraction to sodium dodecyl sulfate results in an "apparent" enhancement of the P700 to chlorophyll ratio to one P700 per 51 light harvesting chlorophyll. Room temperature electron spin resonance measurements of photooxidized F60 are consistent with the presence of P700 and with the chlorophyll/P700 ratio observed by chemical assay. The amino acid compositions of F60 and F65 complexes are studied. Gel electrophoresis patterns of these six isolated complexes are presented and are significantly different from those reported for detergent-treated chlorophyll-protein complexes.

An ultracentrifuge study of C-phycocyanin aggregation.

The molecular weight of C-phycocyanin has been determined as a function of protein concentration in a sodium acetate buffer solution of pH 4.8 at 21.2 degrees C by the Yphantis method. The higher aggregate, the dodecamer (19 S), is first removed by dissociation simply by dialysis against pH 3.9 acetate buffer solution. The molecular weight data are best interpreted by the simultaneous presence of monomers, trimers, and hexamers. Assuming a monomer K13 in equilibrium trimer K36 in equilibrium hexamer equilibrium system, the equilibrium constants have been calculated, and the values are K13 = 1.4 x 10(12) (L/mol)2 and K36 = 6.1 x 10(5) L/mol. The corresponding free energies for each step have been calculated to be -16.3 kcal/mol (delta G degrees 13) and -7.76 kcal/mol (delta G degrees 36). With these values of K13 and K36, the weight-average molecular weight as a function of concentration is calculated. Satisfactory agreement is obtained between the calculated curve and the experimental data. Sedimentation velocity studies performed with a band-forming centerpiece, on the same protein solutions used for the sedimentation equilibrium studies, confirm the presence of three species: monomers, trimers, and hexamers.

Thermodynamics of carbohydrate-lipid interactions.

Thermodynamic analyses of carbohydrate-lipid interactions were performed by investigating the effects of a series of carbohydrates, including monosaccharides, disaccharides, and trisaccharides, on the phase-transition properties of aqueous dispersions of 1,2-dipalmitoyl phosphatidylcholine (DPPC). The temperature of the lipid's main phase transition from the gel to liquid-crystalline phase is essentially unchanged in the presence of carbohydrate. The change in the free energy (delta G) of the transition is zero when a carbohydrate is added to aqueous dispersions of DPPC, while the enthalpy (delta H) and the entropy of the melting of DPPC are decreased. The thermodynamic information was used to examine carbohydrate-lipid interactions. Such interactions were elucidated according to our knowledge of the specific properties of carbohydrates in aqueous solutions and the previously proposed hydrophobic interaction involving hydrocarbon tails of the lipid in aqueous dispersions.