Effect of heme administration on hemopexin metabolism in the rhesus monkey

Clinical conditions such as hemolytic anemias and certain neuromuscular diseases in which serum hemopexin levels are either increased or decreased were simulated in rhesus monkeys by administering heme intravenously daily at three dose levels over a period of 10 days. At the lower dose of heme (0.02 to 0.04 mg/kg/day), serum hemopexin levels were elevated to 150% of control (control = 53.3 +/- 2.8 U/100 ml). At the higher dose of heme (5.0 mg/kg/day), hemopexin levels decreased to 60% of control. After an intermediate dose of heme (0.6 mg/kg/day), no change was seen in the circulating hemopexin levels. These changes appeared to be specific for hemopexin, since neither the serum haptoglobin levels nor the transferrin level was affected by the heme administration at any of the dose levels. Parameters of hemopexin metabolism revealed that after administration of the low dose of heme there was a 76% increase in the net rate of hemopexin synthesis, resulting in a 65% increase in the intravascular pool size of hemopexin. At the intermediate dose there was a 43% increase in the rate of hemopexin synthesis accompanied by a 33% increase in catabolism, resulting in no net change in serum hemopexin concentrations. At the high dose of heme there was a 57% increase in catabolism of hemopexin without a concurrent increase in synthesis, resulting in lowered circulating hemopexin levels. These findings seem to indicate a relationship between the amount of heme presented to the liver and net hemopexin synthesis.     

Hemopexin from four species inhibits the association of heme with cultured hepatoma cells or primary rat hepatocytes exhibiting a small number of species specific hemopexin receptors

Hemopexin (Hx) binds heme with a very high affinity (Kd<0.1 pmol/L). It has been implicated as a major vehicle for the transport of heme into liver cells, involving a receptor-mediated recycling mechanism. However, previous studies indicated that heme is not taken up by cultured embryonic chick or adult rat hepatocytes by such a mechanism, because heme added as heme hemopexin failed to affect heme-responsive activities of 5-aminolevulinic acid synthase and heme oxygenase. Here, we investigated the importance of hemopexin in hepatic heme uptake in cultured rat hepatocytes and human HepG2 hepatoma cells, and determined the number and species specificity of hemopexin receptors on the rat hepatocytes. We also tested whether there is a difference between heterologous and homologous hemopexins. We found the following: 1) heme is inhibited from associating with hepatocytes by apo hemopexins from rat, human, rabbit, and chicken; 2) heme readily associates with hepatocytes when heme hemopexin preparations are added in which the ratio of heme to hemopexin exceeds 1.0; 3) heme induces heme oxygenase mRNA in rat hepatocytes and this induction is prevented by excess hemopexin; and 4) rat hepatocytes exhibit only about 2,000 hemopexin receptors per cell when using rat hemopexin, and none when using hemopexin of rabbit and human. We conclude that hemopexin plays a limited role in heme uptake by cultured hepatocytes and hepatoma cells, and that heme which exceeds the hemopexin binding capacity is taken up directly from heme-albumin.     

Photodissociation of ligands from heme and heme proteins. Effect of temperature and organic phosphate

The effect of temperature on ligand photodissociation from protoheme and the heme proteins hemoglobin (Hb) and myoglobin (Mb) has been examined. The quantum yield of photodissociation (phi) is greater at 40 degrees than at 0 degrees; in general, larger increases are seen in the less photosensitive complexes, while phi does not change in the most photosensitive complexes. The ratio of phi at 40 degrees to phi at 0 degrees is 1.8 for HbCO, 2.3 for n-butyl isocyanide Hb, 2.7 for HbO2, and 1.3 for HbNO, with initial phi values of 0.38, 0.26, 0.028, and 0.003, respectively. This pattern of quantum yield increases is seen in protoheme as well as Hb and Mb ligand photolysis. The allosteric effector inositol hexaphosphate increases the quantum yield of lignad photolysis from hemoglobin. As with temperature, inositol hexaphosphate addition has a larger effect on complexes of low quantum yield; phi increases 1.2-fold for HbCO and 2.2-fold for HbO2 at 0 degrees. The results are discussed in terms of a model containing a photoexcited intermediate (Phillipson, P.E., Ackerson, B.J., and Wyman, J. (1973) Proc. Natl. Acad. Sci. U.S.A. 70, 1550-1553).     

Neisseria gonorrhoeae heme biosynthetic mutants utilize heme and hemoglobin as a heme source but fail to grow within epithelial cells

Many bacterial pathogens, including pathogenic neisseriae, can use heme as an iron source for growth. To study heme utilization by Neisseria gonorrhoeae, two heme biosynthetic mutants were constructed, one with a mutation in hemH (the gene encoding ferrochelatase) and one with a mutation in hemA (the gene encoding gamma-glutamyl tRNA reductase). The hemH mutant failed to grow without an exogenous supply of heme or hemoglobin, whereas the hemA mutant failed to grow unless heme, hemoglobin, or heme precursors were present. Growth of the mutants with hemoglobin required expression of the hemoglobin receptor (HpuAB) and was TonB dependent. However, growth with heme required neither HpuAB nor TonB. An fbpA mutant grew normally when either heme or hemoglobin was present in the medium. The heme biosynthetic mutants showed reduced intracellular survival, compared to the parent strain, within A-431 endocervical epithelial cell cultures. These studies demonstrate that in addition to synthesizing their own heme, N. gonorrhoeae strains are able to internalize and utilize exogenous heme independently of FbpA but appear unable to obtain heme from within epithelial cells for growth.     

Relating matrix metalloproteinase structure to function: why the "hemopexin" domain?

Matrix metalloproteinases are thought to be key players in the remodelling activity of cells associated with both physiological and pathological processes. They share a relatively conserved structure with a number of identifiable modules linked to their specific functions. The structure of the individual domains of a number of matrix metalloproteinases have now been elucidated. Here we review these data in the light of complementary studies on the behaviour of these enzymes in biological systems. In particular we focus on the C-terminal hemopexin-like domain which has intriguingly specific roles in individual matrix metalloproteinases.     

De novo heme proteins from designed combinatorial libraries

We previously reported the design of a library of de novo amino acid sequences targeted to fold into four-helix bundles. The design of these sequences was based on a "binary code" strategy, in which the patterning of polar and nonpolar amino acids is specified explicitly, but the exact identities of the side chains is varied extensively (Kamtekar S, Schiffer JM, Xiong H, Babik JM, Hecht MH, 1993, Science 262:1680-1685). Because of this variability, the resulting collection of amino acid sequences may include de novo proteins capable of binding biologically important cofactors. To probe for such binding, the de novo sequences were screened for their ability to bind the heme cofactor. Among an initial collection of 30 binary code sequences, 15 are shown to bind heme and form bright red complexes. Characterization of several of these de novo heme proteins demonstrated that their absorption spectra and resonance Raman spectra resemble those of natural cytochromes. Because the design of these sequences is based on global features of polar/ nonpolar patterning, the finding that half of them bind heme highlights the power of the binary code strategy, and demonstrates that isolating de novo heme proteins does not require explicit design of the cofactor binding site. Because bound heme plays a key role in the functions of many natural proteins, these results suggest that binary code sequences may serve as initial prototypes for the development of large collections of functionally active de novo proteins.     

Energy Transfer from PVK to Europium Complex in Electroluminescence

The spectra of solutions, films and light-emitting diodes of rare earth complexes were studied. It is shown that the absorption spectra of PVK-dopping rare earth complexes can be red-shifted to the visible region and overlap with the emission spectrum of PVK, which makes the energy transfer possible from PVK to the rare earth ion.     

Transfer of training from reaction time to tachistoscopic recognition.

Describes 2 experiments in which RT practice preceded a tachistoscopic recognition task. Ss were 108 and 180 undergraduates, respectively. The amount of positive transfer obtained was shown to be a function of both the type of RT task and the amount of RT practice. This transfer was attributed to the learning of an attentional response to the foreperiod which was the same length in both tasks. (French summary) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The heme prosthetic group of lactoperoxidase. Structural characteristics of heme l and heme l-peptides

The heme prosthetic group from the bovine milk enzyme lactoperoxidase (LPO), termed heme l, is isolated through an approach that combines proteolytic hydrolysis and reverse-phase high performance liquid chromatographic separation of the resulting digest. Application of different proteases yields either a peptide-bound heme (with trypsin and chymotrypsin) or a peptide-free heme (with proteinase K). Both heme l and heme l-peptide species were investigated by paramagnetic 1H NMR spectroscopy, electrospray mass spectrometry, and peptide sequence analysis. Paramagnetic 1H NMR experiments on the low spin bis(cyano)-Fe(III)heme l complex conclusively define the heme l structure as a 1,5-bis(hydroxymethyl) derivative of heme b. The electrospray mass spectrum of heme l confirms the two-site hydroxyl functionalization on this heme. Paramagnetic 1H NMR spectra of the high spin bis(dimethyl sulfoxide)-Fe(III) complexes of the isolated heme species provide information regarding peptide content. Sequence analyses of peptides released from two heme l-peptide species by base hydrolysis suggest that heme-protein ester linkages in lactoperoxidase occur between the two hydroxyl groups of heme l and the carboxylic side chains of glutamate 275 and aspartate 125. These results confirm the earlier reported structural proposal (Rae, T. D., and Goff, H. M. (1996) J. Am. Chem. Soc. 118, 2103-2104).     

Transfer from conceptual rule problems to a truth-table-sorting task.

Tested the proposal that during conceptual rule learning, Ss encode the stimulus population into 4 stimulus subclasses which are analogous to the categories of a logical truth table. In Exp I (n = 48), 1st and 2nd graders were given 2 or 4 conceptual problems to solve, the number of rules within the series of problems being either 1 or 2 (conjunction or inclusive disjunction). 2 groups were included to control the effects of warm-up and generalized learning sets. Analysis of errors made on the sorting task showed the number of rules that define solution to be a reliable source of variation. Exp II with 48 1st and 2nd graders and 48 undergraduates was similar to Exp I. Number of rules and the interaction of number of rules and age of S were significant variables. Results are taken as support for the stimulus-encoding hypothesis in modified form. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Dynamic Axial Load Transfer from Elastic Bar to Poroelastic Medium

The time-harmonic response of a cylindrical elastic bar (pile) partially embedded in a homogeneous poroelastic medium and subjected to a vertical load is considered. The bar is modeled using 1D elastic theory valid for long bars in the low-frequency range, and the porous medium using Biot's 3D elastodynamic theory. The bar is bonded to the surrounding medium along the contact surface. The problem is formulated by decomposing the bar∕porous medium system into a fictitious bar and an extended porous medium. A Fredholm's integral equation of the second kind governs the distribution of axial force in the fictitious bar. The integral equation involves kernels that are displacement and strain influence functions of a poroelastic half-space subjected to a buried, uniform vertical patch load. The governing integral equation is solved by applying numerical quadrature. The solutions for axial displacement and axial force of the bar, and the pore pressure are also derived. Selected numerical results for vertical impedance, axial force, and pore pressure profiles are presented to portray the influence of bar stiffness and length∕radius ratio, frequency of excitation, and poroelastic properties.     

Transfer of value from fit.

People experience regulatory fit when they pursue a goal in a manner that sustains their regulatory orientation (E. T. Higgins, 2000). Five studies tested whether the value experienced from regulatory fit can transfer to a subsequent evaluation of an object. In Studies 1 and 2, participants gave the same coffee mug a higher price if they had chosen it with a strategy that fit their orientation (eager strategy/promotion; vigilant strategy/prevention) than a strategy that did not fit. Studies 3-5 investigated possible mechanisms underlying this effect. Value transfer was independent of positive mood, perceived effectiveness (instrumentality), and perceived efficiency (ease), and occurred for an object that was independent of the fit process itself. The findings supported a value confusion account of transfer. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Transfer of value from S+ to S- in a simultaneous discrimination.

Value transfer theory has been proposed by L. von Fersen et al (see record 1991-29523-001) to account for transitive inference effects, in which following training on 4 simultaneous discriminations (A?+?B–, B?+?C–, C?+?D–, D?+?E–) pigeons showed a preference for B over D. According to this theory, some of the value of reinforcement acquired by each stimulus always associated with reinforcement (S+) transfers to the stimulus associated with nonreinforcement (S–). In the transitive inference experiment, C (associated with both reward and nonreward) could transfer less value to D than A (associated only with reward) could transfer to B. Support for value transfer theory was demonstrated in 2 experiments, involving a total of 20 pigeons, in which an S– presented in the context of an S+ was preferred over an S– presented in the context of a stimulus to which responses were sometimes reinforced (S±). (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Identification of histidine 45 as the axial heme iron ligand of heme oxygenase-2

A truncated, soluble, and enzymatically active form of human heme oxygenase-2 (DeltaHHO2) was expressed in Escherichia coli. To identify the axial heme ligand of HO-2, His-45 to Ala (DeltaH45A) and His-152 to Ala (DeltaH152A) mutants have been prepared using this expression system. DeltaH45A could form a 1:1 complex with hemin but was completely devoid of the heme degradation activity. A 5-coordinate-type ferrous NO EPR spectrum was observed for the heme-DeltaH45A complex. The DeltaH152A mutant was expressed as an inclusion body and was recovered from the lysis pellet by dissolution in urea followed by dialysis. The solubilized fraction obtained, however, was composed of a mixture of a functional enzyme and an inactive fraction. The inactive fraction was removed by Sephadex G-75 column chromatography since it eluted out of the column at the void volume. The gel filtration-purified DeltaH152A exhibited spectroscopic and enzymatic properties identical to those of wild-type. We conclude, in contrast to the previous reports (McCoubrey and Maines (1993) Arch. Biochem. Biophys. 302, 402-408; McCoubrey, W. K., Jr., Huang, T. J., and Maines, M. (1997) J. Biol. Chem. 272, 12568-12574), that His-45, but not His-152, in heme oxygenase isoform-2 is the proximal ligand of the heme and is essential for the heme degradation activity of the enzyme. His-152 appears to play a structural role in stabilization of the heme oxygenase protein.     

Selenium regulation of hepatic heme metabolism: induction of delta-aminolevulinate synthase and heme oxygenase

Selenium was found to be a novel regulator of cellular heme methabolism in that the element induced both the mitochondrial enzyme delta-aminolevulinate synthase [succinyl-CoA:glycine C-succinyltransferase (decarboxylating); EC 2-3-1-37] and the microsomal enzyme heme oxygenase [heme, hydrogen-donor:oxygen oxidoreductase(alpha-methene-oxidizing, hydroxylating); EC 1-14-99-3] in liver. The effect of selenium on these enzyme activities was prompt, reaching a maximum within 2 hr after a single injection. Other changes in parameters of hepatic heme metabolism occurred after administration of the element. Thirty minutes after injection the cellular content of heme was significantly increased; however, this value slightly decreased below control values within 2 hr, coinciding with the period of rapid induction of heme oxygenase. At later peroids heme content returned to normal values. Selenium treatment caused only a slight decrease in microsomal cytochrome P-450 content. However, drug-metabolizing activity was severely inhibited by higher doses of the element. Unlike other inducers of delta-aminolevulinate synthase, which as a rule are also porphyrinogenic agents, selenium induction of this enzyme was not accompanied by an increase in the cellular content of prophyrins. When rats were pretreated with selenium 90 min before administration of heme, a potent inhibitor of delta-aminolevulinate synthase production, the inhibitory effect of heme of formation of this mitochondrial enzyme was completely blocked. Selenium, at high concentrations in vitro, was inhibitory to delta-aminolevulinate synthase activity. It is postulated that selenium may not be a direct inducer of heme oxygenase as is the case with trace metals such as cobalt, but may mediate an increase in heme oxygenase through increased production and cellular availability of "free" heme, which results from the increased heme synthetic activity of hematocytes. Subsequently, the increased heme oxygenase activity is in turn responsible for the lack of increase in the microsomal heme content, thus maintaining heme levels at normal values despite the highly increased activities of both heme oxygenase and delta-aminolevulinate synthase. It is further suggested that the increase in delta-aminolevulinate synthase activity is not due to a decreased rate of enzyme degradation or an activation of preformed enzyme, but to increased rate of synthesis of enzyme protein. Although selenium in trace amounts has been postulated to be involved in microsomal electron transfer process, the data from this study indicate that excess selenium can substantially inhibit microsomal drug metabolism.     

Molecular mechanism of heme biosynthesis

Two of the major organs producing heme are bone marrow and the liver. delta-Aminolevulinate synthase (ALAS) plays the key role to regulate heme biosynthesis in hepatocytes as well as in erythroid cells. In the liver, nonspecific (or housekeeping) isozyme of ALAS (ALAS-N) is expressed to be regulated by its end product, heme, in a negative feedback manner. The way to regulate ALAS-N in the liver is suitable to supply a constant level of heme for a family of drug metabolizing enzymes, cytochrome P-450 (CYP). In erythroid tissues, not only erythroid-specific isozyme of ALAS (ALAS-E) but also ALAS-N are expressed, and regulated by distinctive manners. Although heme regulates ALAS-N in a negative feedback manner even in erythroid cells, ALAS-E is upregulated by induced heme concentration. ALAS-N in undifferentiated erythroid cells, therefore, is suggested to produce heme for CYP, whereas heme for accumulating hemoglobin (Hb) in cells undergoing differentiation is synthesized via ALAS-E. In this article, we describe the molecular mechanisms to regulate heme biosynthesis in non-erythroid as well as in erythroid tissues, and discuss the pathological significance of the mechanisms in patients with inherited disorders, porphyrias.     

Transfer effects of success and failure training from one reinforcing agent to another.

Determined the transfer effects of behavior changes in the presence of 1 reinforcing agent (Experimenter A) on behavior in the presence of another agent (Experimenter B). Ss were 36 boys 6-10 yrs old in grades 1-3, judged to have poor attention span and low achievement in arithmetic. The experimental task consisted of a series of counting problems. Each session included 2 12-min subsessions during which S could earn token reinforcement only by attending to the task and solving problems. Experimenter A conducted 1 subsession and Experimenter B conducted the other. An O recorded inappropriate classroom behavior (talking and out-of-seat behavior). Following the establishment of stable, intermediate baseline levels of counting, Experimenter A introduced a more favorable reinforcement schedule (success training) for some Ss and a less favorable schedule (failure training) for some Ss, but continued baseline schedules for Ss in the control condition. Experimenter B continued baseline schedules for all Ss. The effects of success training produced generalization of greater counting behavior while the effects of failure training produced no transfer of less counting behavior and generalization of greater inappropriate classroom behavior. There was no evidence for behavior contrast. (33 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)