Synthesis of a metal binding protein designed on the alpha/beta scaffold of charybdotoxin

The alpha/beta scaffold of the scorpion toxin charybdotoxin has been used for the engineering of a metal binding site. Nine substitutions, including three histidines as metal ligands, have been introduced into the original toxin sequence. The newly designed sequence, 37 amino acids long, has been assembled by solid-phase synthesis and HBTU (2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate) coupling of Fmoc-protected amino acids. Formation of the three disulfide bonds occurred efficiently and rapidly in the presence of glutathione, and this post-synthesis modification has facilitated the purification task enormously. The process of synthesis and purification was performed in less than a week with an overall 10.2% yield. Circular dichroism analysis showed that the newly designed protein is folded in a alpha/beta structure, similarly to the parent toxin. Electronic absorption spectroscopy, circular dichroism and gel filtration experiments have been used to show that Cu2+ and Zn2+ ions bind with high affinity to the newly engineered protein. These results demonstrate that the alpha/beta fold, common to all scorpion toxins, is a very versatile basic structure, tolerant for substitutions and able to present new sequences in a predetermined conformation. The chemical approach is shown to be effective, rapid and practical for the production of novel designed small proteins.     

NMR solution structure of a two-disulfide derivative of charybdotoxin: structural evidence for conservation of scorpion toxin alpha/beta motif and its hydrophobic side chain packing

The alpha/beta scorpion fold consisting of a short alpha-helix and beta-sheet is a structural motif common to scorpion toxins, insect defensins, and plant gamma-thionins that invariably contains three disulfides. CHABII is a two-disulfide derivative of the scorpion toxin charybdotoxin (ChTX), chemically synthesized by inserting two L-alpha-aminobutyric acids in place of the two half-cystine residues involved in the disulfide 13-33. This disulfide is one of the two disulfides which connect the alpha-helix to the beta-sheet. The solution structure of CHABII was determined at pH 6.3 and 5 degrees C using 2D NMR and simulated annealing from 513 distance and 46 dihedral angle constraints. The NMR structure of CHABII is well-defined as judged from the low value of the averaged backbone rms deviation between the 30 lowest energy structures and the energy-minimized mean structure ((rmsd) = 0.65 A for the entire sequence and 0.48 A for the segment 3-36). Analysis and comparison of the solution structures of CHABII and ChTX lead to the following conclusions: (i) the fold of CHABII is similar to that of ChTX as indicated by the low value of the averaged backbone atomic rms deviation between the 10 lowest energy solution structures of the two proteins (1.44 A); (ii) the packing of the hydrophobic core is well-preserved, underlying the critical structural role of the hydrophobic interactions even for such a small and cysteine-rich protein as ChTX.     

Biochemical characterization of human osteoclast integrins. Osteoclasts express alpha v beta 3, alpha 2 beta 1, and alpha v beta 1 integrins

The study of osteoclast integrins has been previously hampered by the lack of a source of large numbers of purified osteoclasts. Osteoclastoma, a human giant cell tumor of bone, supplied a rich source of osteoclasts within a tissue containing many diverse cell types. Osteoclastoma integrin immunostaining confirmed the presence of the integrin alpha v beta 3 complex and the alpha 2 and beta 1 integrin subunits on osteoclasts. However, weak integrin expression, for example with alpha v beta 5, was difficult to interpret. Purification with magnetic beads coated with vitronectin receptor monoclonal antibody (13C2) enabled osteoclast membranes to be isolated with high purity and yield (57%) from osteoclastoma tissue. Positively (osteoclast-enriched) selected membranes were biochemically assessed for integrin expression by immunoprecipitation and visualization by non-radioactive enhanced chemiluminescence. alpha 1, alpha 4, alpha 6, alpha 8, alpha M, alpha X, gpIIb, beta 4, beta 6, and beta 8 integrin chains were undetectable at a sensitivity of 1 ng. alpha 3, alpha 5, alpha L, beta 2, and alpha v beta 5 were found in the negatively selected osteoclastoma tissue but not in the positively purified osteoclast membranes. The presence of alpha v beta 1 and alpha 2 beta 1 dimers was demonstrated biochemically on the immunoisolated osteoclast membranes. Osteoclast alpha v beta 3 isolation by Arg-Gly-Asp (RGD) affinity chromatography for NH2-terminal amino acid sequencing confirmed that the osteoclast vitronectin receptor was identical to that previously characterized on other cell types. In situ hybridization using human alpha v riboprobes in osteoclasts from human and rodent bone further demonstrated the high level and specificity of expression of alpha v vitronectin receptor in osteoclasts.     

NMR structures and activity of a novel alpha-like toxin from the scorpion Leiurus quinquestriatus hebraeus

NMR structures of a new toxin from the scorpion Leiurus quinquestriatus hebraeus (Lqh III) have been investigated in conjunction with its pharmacological properties. This toxin is proposed to belong to a new group of scorpion toxins, the alpha-like toxins that target voltage-gated sodium channels with specific properties compared with the classical alpha-scorpion toxins. Electrophysiological analysis showed that Lqh III inhibits a sodium current inactivation in the cockroach axon, but induces in addition a resting depolarization due to a slowly decaying tail current atypical to other alpha-toxin action. Binding studies indicated that radiolabeled Lqh III binds with a high degree of affinity (Ki=2.2 nM) on cockroach sodium channels and that the alpha-toxin from L quinquestriatus hebraeus highly active on insects (LqhalphaIT) and alpha-like toxins compete at low concentration for its receptor binding site, suggesting that the alpha-like toxin receptor site is partially overlapping with the receptor site 3. Conversely, in rat brain, Lqh III competes for binding of the most potent anti-mammal alpha-toxin from Androctonus australis Hector venom (AaH II) only at very high concentration. The NMR structures were used for the scrutiny of the similarities and differences with representative scorpion alpha-toxins targeting the voltage-gated sodium channels of either mammals or insects. Three turn regions involved in the functional binding site of the anti-insect LqhalphaIT toxin reveal significant differences in the Lqh III structure. The electrostatic charge distribution in the Lqh III toxin is also surprisingly different when compared with the anti-mammal alpha-toxin AaH II. Similarities in the electrostatic charge distribution are, however, recognized between alpha-toxins highly active on insects and the alpha-like toxin Lqh III. This affords additional important elements to the definition of the new alpha-like group of scorpion toxins and the mammal versus insect scorpion toxin selectivities.     

Treatment of snake, insect, scorpion, and spider bites in the pediatric emergency department

Obtaining an adequate depth of anesthesia is a continuous challenge to the anesthetist. With the introduction of muscle-relaxing agents the traditional signs of awareness are often obscured, or difficult to interpret. These signs include blood pressure, heart rate, pupil size, etc. However, these factors do not describe the depth of anesthesia (DA) in a cerebral activity sense. Hence, a better measure of the DA is required. It has been suggested that Auditory-Evoked Potentials (AEP) can provide additional information about the DA. The general method of extracting AEP is by use of a Moving Time Average (MTA). However, the MTA is time consuming because a large number of repetitions is needed to produce an estimate of the AEP. Hence, changes occurring over a small number of sweeps will not be detected by the MTA average. We describe a system-identification method, an autoregressive model with exogeneous input (ARX) model, to produce a sweep-by-sweep estimate of the AEP. The method was clinically evaluated in 10 patients anesthetized with alfentanil and propofol. The time interval between propofol induction and the time when the Na-Pa amplitude was decreased to 25% of the initial amplitude was measured. These measurements showed that ARX-estimated compared to MTA-estimated AEP was significantly faster in tracing transition from consciousness to unconsciousness during propofol induction (p < 0.05).     

An excitatory scorpion toxin with a distinctive feature: an additional alpha helix at the C terminus and its implications for interaction with insect sodium channels

BACKGROUND: Scorpion neurotoxins, which bind and modulate sodium channels, have been divided into two groups, the alpha and beta toxins, according to their activities. The beta-toxin class includes the groups of excitatory and depressant toxins, which differ in their mode of action and are highly specific against insects. The three-dimensional structures of several alpha and beta toxins have been determined at high resolution, but no detailed 3D structure of an excitatory toxin has been presented so far. RESULTS: The crystal structure of an anti-insect excitatory toxin from the scorpion Buthotus judaicus, Bj-xtrIT, has been determined at 2.1 A resolution and refined to an R factor of 0.209. The first 59 residues form a closely packed module, structurally similar to the conserved alpha and beta toxins ('long toxins') affecting sodium channels. The last 17 residues form a C-terminal extension not previously seen in scorpion toxins. It comprises a short alpha helix anchored to the N-terminal module by a disulfide bridge and is followed by a highly mobile stretch of seven residues, of which only four are seen in the electron-density map. This mobile peptide covers part of a conserved hydrophobic surface that is thought to be essential for interaction with the channel in several long toxins. CONCLUSIONS: Replacement of the last seven residues by a single glycine abolishes the activity of Bj-xtrIT, strongly suggesting that these residues are intimately involved in the interaction with the channel. Taken together with the partial shielding of the conserved hydrophobic surface and the proximity of the C terminus to an adjacent surface rich in charged residues, it seems likely that the bioactive surface of Bj-xtrIT is formed by residues surrounding the C terminus. The 3D structure and a recently developed expression system for Bj-xtrIT pave the way for identifying the structural determinants involved in the bioactivity and anti-insect specificity of excitatory toxins.     

Differences in interferon alpha and beta signaling. Interferon beta selectively induces the interaction of the alpha and betaL subunits of the type I interferon receptor

All Type I interferons (IFNalpha, IFNbeta, IFNomega) bind to the Type I IFN receptor (IFNR) and elicit a common set of signaling events, including activation of the Jak/Stat and IRS pathways. However, IFNbeta selectively induces the association of the alpha subunit of the Type I IFNR with p100, a tyrosyl phosphoprotein, to transduce IFNbeta-specific signals. Using antibodies raised against the different components of the Type I IFNR, we identified p100 as the long form of the beta subunit (betaL subunit) of the Type I IFNR. This was also confirmed in experiments with mouse L-929 cells transfected with truncated forms of betaL. Thus, IFNbeta stimulation of human cells or mouse L-929 transfectants expressing the human alpha and betaL subunits, selectively induces the formation of a signaling complex containing the alpha and betaL subunits of the receptor. The IFNbeta-regulated interaction of the alpha and betaL chains is rapid and transient and follows a similar time course with the tyrosine phosphorylation of these receptor components. These data demonstrate that the signaling specificity for different Type I IFNs is established early in the signaling cascade, at the receptor level, and results from distinct interactions between components of the Type I IFNR.     

Molecular determinants of high affinity binding of alpha-scorpion toxin and sea anemone toxin in the S3-S4 extracellular loop in domain IV of the Na+ channel alpha subunit

alpha-Scorpion toxins and sea anemone toxins bind to a common extracellular site on the Na+ channel and inhibit fast inactivation. Basic amino acids of the toxins and domains I and IV of the Na+ channel alpha subunit have been previously implicated in toxin binding. To identify acidic residues required for toxin binding, extracellular acidic amino acids in domains I and IV of the type IIa Na+ channel alpha subunit were converted to neutral or basic amino acids using site-directed mutagenesis, and altered channels were transiently expressed in tsA-201 cells and tested for 125I-alpha-scorpion toxin binding. Conversion of Glu1613 at the extracellular end of transmembrane segment IVS3 to Arg or His blocked measurable alpha-scorpion toxin binding, but did not affect the level of expression or saxitoxin binding affinity. Conversion of individual residues in the IVS3-S4 extracellular loop to differently charged residues or to Ala identified seven additional residues whose mutation caused significant effects on binding of alpha-scorpion toxin or sea anemone toxin. Moreover, chimeric Na+ channels in which amino acid residues at the extracellular end of segment IVS3 of the alpha subunit of cardiac Na+ channels were substituted into the type IIa channel sequence had reduced affinity for alpha-scorpion toxin characteristic of cardiac Na+ channels. Electrophysiological analysis showed that E1613R has 62- and 82-fold lower affinities for alpha-scorpion and sea anemone toxins, respectively. Dissociation of alpha-scorpion toxin is substantially accelerated at all potentials compared to wild-type channels. alpha-Scorpion toxin binding to wild type and E1613R had similar voltage dependence, which was slightly more positive and steeper than the voltage dependence of steady-state inactivation. These results indicate that nonidentical amino acids of the IVS3-S4 loop participate in alpha-scorpion toxin and sea anemone toxin binding to overlapping sites and that neighboring amino acid residues in the IVS3 segment contribute to the difference in alpha-scorpion toxin binding affinity between cardiac and neuronal Na+ channels. The results also support the hypothesis that this region of the Na+ channel is important for coupling channel activation to fast inactivation.     

Stem-cell-associated structural and functional plasticity in the aging hippocampus.

Aging frequently leads to a functional decline across multiple cognitive domains, often resulting in a severe reduction in life quality and also causing substantial care-related costs. Understanding age-associated structural and functional changes of neural circuitries within the brain is required to improve successful aging. In this review, the authors focus on age-dependent alterations of the hippocampus and the decline of hippocampal function, which are critically involved in processes underlying certain forms of learning and memory. Despite the dramatic reductions in hippocampus-dependent function that accompany advancing age, there is also striking evidence that even the aged brain retains a high level of plasticity. Thus, one promising avenue to reach the goal of successful aging might be to boost and recruit this plasticity, which is the interplay between neural structure, function, and experience, to prevent age-related cognitive decline and age-associated comorbidities. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Synthesis and conformational studies of peptides containing TOAC, a spin-labelled C alpha, alpha-disubstituted glycine

A variety of host L-alanine homo-peptides (to the pentamer) containing one or two spin-labelled TOAC (2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid) residues were synthesized by solution methods and fully characterized. The conformational features of the terminally blocked, doubly spin-labelled TOAC-(Ala)2-TOAC-Ala-pentapeptide were examined in the crystal state by X-ray diffraction and in solution using a combination of techniques (Fourier transform infrared, circular dichroism, cyclic voltammetry and electron spin resonance) in comparison with singly labelled shorter peptides. The 3(10)-helical structure of the pentapeptide, promoted by the two C alpha, alpha-disubstituted glycines under favourable experimental conditions, allows an interaction to take place between the two nitroxide TOAC side chains spaced by one turn of the helix. Taken together, these results suggest that TOAC is an excellent probe for exploring bends and helices in doubly labelled peptides.     

Sequence and functional characterization of a renal sodium/dicarboxylate cotransporter

The cDNA coding for a rabbit renal Na+/dicarboxylate cotransporter, designated NaDC-1, was isolated by functional expression in Xenopus oocytes. NaDC-1 cDNA is approximately 2.3 kilobases in length and codes for a protein of 593 amino acids. NaDC-1 protein contains eight putative transmembrane domains, and the sequence and secondary structure are related to the renal Na+/sulfate transporter, NaSi-1. Northern analysis shows that the NaDC-1 message is abundant in kidney and small intestine, and related transporters may be found in liver, lung, and adrenal. The transport of succinate by NaDC-1 was sodium-dependent, sensitive to inhibition by lithium, and inhibited by a range of di- and tricarboxylic acids. This transporter also carries citrate, but it does not transport lactate. In kinetic experiments, the Km for succinate was around 0.4 mM and the Vmax was 15 nmol/oocyte/h, while the Hill coefficient of Na+ activation of succinate transport was 1.9. The transport of succinate by NaDC-1 was insensitive to changes in pH, whereas the transport of citrate increased with decreasing pH, in parallel with the concentration of divalent citrate in the medium. The results of the functional characterization indicate that NaDC-1 likely corresponds to the renal brush-border Na+/dicarboxylate cotransporter.     

Differential interactions of the C terminus and the cytoplasmic I-II loop of neuronal Ca2+ channels with G-protein alpha and beta gamma subunits. I. Molecular determination

Interactions of G-protein alpha (Galpha) and beta gamma subunits (Gbeta gamma) with N- (alpha1B) and P/Q-type (alpha1A) Ca2+ channels were investigated using the Xenopus oocyte expression system. Gi3alpha was found to inhibit both N- and P/Q-type channels by receptor agonists, whereas Gbeta1 gamma2 was responsible for prepulse facilitation of N-type channels. L-type channels (alpha1C) were not regulated by Galpha or Gbeta gamma. For N-type, prepulse facilitation mediated via Gbeta gamma was impaired when the cytoplasmic I-II loop (loop 1) was deleted or replaced with the alpha1C loop 1. Galpha-mediated inhibitions were also impaired by substitution of the alpha1C loop 1, but only when the C terminus was deleted. For P/Q-type, by contrast, deletion of the C terminus alone diminished Galpha-mediated inhibition. Moreover, a chimera of L-type with the alpha1B loop 1 gained Gbeta gamma-dependent facilitation, whereas an L-type chimera with the N- or P/Q-type C terminus gained Galpha-mediated inhibition. These findings provide evidence that loop 1 of N-type channels is a regulatory site for Gbeta gamma and the C termini of P/Q- and N-types for Galpha.     

Working in a majority context: A structural model of heterosexism as minority stress in the workplace.

This study used the minority stress theory to develop models of antecedents and outcomes of heterosexism in the workplace. Heterosexism was assessed via the Workplace Heterosexist Experiences Questionnaire, a new scale assessing direct (e.g., anti-gay jokes) and indirect experiences (e.g., assumptions of heterosexuality). Structural equation modeling with two community samples (N?=?287) of gay, lesbian, and bisexual people indicated that heterosexism was best predicted by perceptions that the employer does not take heterosexism seriously (i.e., organizational climate). Experiencing heterosexism was associated with adverse psychological, health, and job-related outcomes, thus supporting the minority stress theory. In addition, outness was positively related to experiences of direct heterosexism but negatively related to indirect experiences. The minority stress theory is discussed in light of the mental health history of pathologizing homosexuality. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Purification and partial characterization of a ''short'' insectotoxin-like peptide from the venom of the scorpion Parabuthus schlechteri

OBJECTIVE: The aim of this paper is to chart the ethical territory surrounding the issue of consensual sexual relationships between psychiatrists and doctors training in psychiatry. METHOD: The arguments for and against the prohibition of such relationships are critically examined both in general and in a number of specific circumstances. RESULTS: There should not be a general prohibition against such relationships, but a prohibition should apply in certain special circumstances. Such circumstances include occasions when the psychiatrist is currently supervising the trainee, when a particular psychiatrist has repeated sexual relationships with trainees and when a group of psychiatrists voluntarily pledge to abstain from such relationships. CONCLUSIONS: Institutions involved in psychiatric training should develop guidelines for dealing with such relationships, adopting a generally permissive attitude toward them with clear exceptions in special cases.     

Differential interactions of the C terminus and the cytoplasmic I-II loop of neuronal Ca2+ channels with G-protein alpha and beta gamma subunits. II. Evidence for direct binding

The present study was designed to obtain evidence for direct interactions of G-protein alpha (Galpha) and beta gamma subunits (Gbeta gamma) with N- (alpha1B) and P/Q-type (alpha1A) Ca2+ channels, using synthetic peptides and fusion proteins derived from loop 1 (cytoplasmic loop between repeat I and II) and the C terminus of these channels. For N-type, prepulse facilitation as mediated by Gbeta gamma was impaired when a synthetic loop 1 peptide was applied intracellularly. Receptor agonist-induced inhibition of N-type as mediated by Galpha was also impaired by the loop 1 peptide but only when applied in combination with a C-terminal peptide. For P/Q-type channels, by contrast, the Galpha-mediated inhibition was diminished by application of a C-terminal peptide alone. Moreover, in vitro binding analysis for N- and P/Q-type channels revealed direct interaction of Galpha with C-terminal fusion proteins as well as direct interaction of Gbeta gamma with loop 1 fusion proteins. These findings define loop 1 of N- and P/Q-type Ca2+ channels as an interaction site for Gbeta gamma and the C termini for Galpha.     

Synthesis and characterization of the structural and electrochemical properties of Nd-Al codoped amorphous nickel hydroxide

Nd-Al codoped amorphous nickel hydroxide powders were synthesized by microemulsion precipitation method combined with rapid freezing technique.The microstructure of the prepared samples was analyzed with X-ray diffractometer(XRD),scanning electron microscopy(SEM),and Raman spectroscopy.The electrochemical performances of the prepared samples were characterized with charge/discharge test,cyclic voltammetry,and electrochemical impedance spectra.The results showed that the codoping of Nd-Al resulted in more st...     

The gamma 2 subunit of the GABAA receptor is concentrated in synaptic junctions containing the alpha 1 and beta 2/3 subunits in hippocampus, cerebellum and globus pallidus

The gamma 2 subunit is necessary for the expression of the full benzodiazepine pharmacology of GABAA receptors and is one of the major subunits in the brain. In order to determine the location of channels containing the gamma 2 subunit in relation to GABA-releasing terminals on the surface of neurons, a new polyclonal antipeptide antiserum was developed to the gamma 2 subunit and used in high resolution, postembedding, immunoelectron-microscopic procedures. Dual immunogold labelling of the same section for two subunits, and up to three sections of the same synapse reacted for different subunits, were used to characterize the subunit composition of synaptic receptors. The gamma 2 subunit was present in type 2, "symmetrical" synapses in each of the brain areas studied, with the exception of the granule cell layer of the cerebellum. The gamma 2 subunit was frequently co-localized in the same synaptic junction with the alpha 1 and beta 2/3 subunits. The immunolabelling of synapses was coincident with the junctional membrane specialization of the active zone. Immunolabelling for the receptor often occurred in multiple clusters in the synapses. In the hippocampus, the gamma 2 subunit was present in basket cell synapses on the somata and proximal dendrites and in axo-axonic cell synapses on the axon initial segment of pyramidal and granule cells. Some synapses on the dendrites of GABAergic interneurones were densely labelled for the gamma 2, alpha 1 and beta 2/3 subunits. In the cerebellum, the gamma 2 subunit was present in both distal and proximal Purkinje cell dendritic synapses established by stellate and basket cell, respectively. On the soma of Purkinje cells, basket cell synapses were only weakly labelled. Synapses on interneuron dendrites were more densely labelled for the gamma 2, alpha 1 and beta 2/3 subunits than synapses on Purkinje or granule cells. Although immunoperoxidase and immunofluorescence methods show an abundance of the gamma 2 subunit in granule cells, the labelling of Golgi synapses was much weaker with the immunogold method than that of the other cell types. In the globus pallidus, many type 2 synapses were labelled for the gamma 2 subunit together with alpha 1 and beta 2/3 subunits. The results show that gamma 2 and beta 2/3 subunits receptor channels are highly concentrated in GABAergic synapses that also contain the alpha 1 and beta 2/3 subunits. Channels containing the gamma 2 subunit are expressed in synapses on functionally distinct domains of the same neuron receiving GABA from different presynaptic sources. There are quantitative differences in the density of GABAA receptors at synapses on different cell types in the same brain area.     

Use of Vibrio spp. for expression of Escherichia coli enterotoxin B subunit fusion proteins: purification and characterization of a chimera containing a C-terminal fragment of DNA polymerase from herpes simplex virus type 1

The nontoxic B subunit of Escherichia coli heat-labile enterotoxin (EtxB) is a convenient carrier molecule for the attachment and delivery of heterologous peptides into eukaryotic cells. To evaluate the properties of such EtxB-based fusion proteins an efficient method for their production and purification is required. High-level production and purification of native EtxB has been achieved using heterologous expression and secretion in a marine Vibrio (Amin, T., and Hirst, T. R., 1994, Protein Expression Purif. 5, 198-204). However, the use of this method to isolate EtxB fusion proteins has been precluded because of their susceptibility to degradation by extracellular proteases secreted by members of the Vibrionaceae. In this paper a method is described for production of EtxB-pol, comprising the enterotoxin B subunit linked to a 27-residue C-terminal fragment of Pol, the catalytic subunit of DNA polymerase of herpes simplex virus type 1 (HSV-1). Following assessment of the relative efficacy of different Vibrio strains as hosts for EtxB-pol expression, the chimera was produced at the highest level of 3.5 mg/liter by cultures of Vibrio sp.60. Addition of 0.3 mM EDTA to the growth medium blocked proteolysis of the secreted EtxB-pol fusion protein, which was then purified to homogeneity using ammonium sulfate fractionation and hydrophobic interaction chromatography, with a yield of 57%. Purified EtxB-pol reacted with both anti-EtxB and anti-Pol peptide antibodies, and was able to specifically bind UL42, a processivity factor which normally binds to the C-terminal region of HSV-1 Pol. This modified method for expression and purification of EtxB-pol should be of general utility for the preparation of other EtxB-based fusion proteins.     

Hb Grady and alpha thalassemia: a contribution to the problem of the number of Hb alpha structural loci in man

Hematological evaluation and data from chain synthesis analyses in six members of the family with two members having Hb Grady (i.e., and alpha chain variant with elongated chains due to an insertion of three amino acid residues [1]) indicate the presence of multiple nonallelic Hb alpha structural loci in the single Hb Grady heterozygote. The data support the earlier stated hypothesis that the Hb alpha Grady locus resulted from a crossing over between chromosomes of two tandemly repeated Hb alpha loci. The presence of an alpha thalassemia condition in one of the two Hb Grady heterozygotes increases the relative production of the alpha Grady chain by a factor of two.