Beta-subunit Variant Research Articles

Characterization of beta subunit variants of recombinant human chorionic gonadotrophin

Human chorionic gonadotropin (hCG), an endogenous glycoprotein hormone, has been widely used for the treatment of infertility and corpus luteum defect in women. The biological specificity of hCG is essentially determined by its beta (β-) subunit, whereas the alpha (α-) subunit is a common subunit shared among the gonadotropin family. In development of a therapeutic recombinant hCG, the purity analysis showed that the beta (β-) subunit has two variants, β1 and β2. Structural characterization using a combination of analytical techniques has demonstrated that β1-subunit is derived from non-glycosylation at Asn 13, whereas β2-subunit is a normal species with complete N-glycosylation at both Asn 13 and Asn 30. In vivo Bioactivity evaluation of the r-hCG fractions with various ratios of β1-and β2-subunits showed that incomplete glycosylation at Asn 13 potentially reduced the biological activity of r-hCG to promote uterus growth. Although hCG has a long history of medicinal use, this is the first report to identify the structural difference of hCG β-subunit variants, as well as to preliminary establish the structure-activity relationship of this variation. The obtained results also suggest the importance of variant characterization and necessary quality control of product variants during the development of recombinant protein therapeutics.

Human chorionic gonadotropin production by a benign lung nodule

Ectopic hormone production by a neoplasm is defined as secretion of a hormone by cells not normally a source of that hormone. This relationship is a recognized phenomenon that assists in diagnosis of disease and provides a marker for monitoring treatments. Ectopic human chorionic gonadotropin (hCG) is associated with neoplasms originating in the lungs, gastrointestinal tract, cervix, and bladder. Recent evidence recognizes 3 variants of hCG, with nontrophoblastic tumors producing the free beta-subunit variant. This variant influences cancer cell growth andmay be valuable as a poor prognostic marker [1]. Approximately 15%of lung tumorsdemonstrate detectable hormone [2], and evidence suggests that paraneoplastic lung tumors are often more resistant to treatment, have a poor prognosis, and are rapidly fatal [3]. This report presents an individual with a benign indolent lung nodule secreting beta-hCG. Final pathology revealed an inflammatory myofibroblastic tumor (IMT). This type of tumor is rare, accounting for 0.4%–1.0% of all lung tumors; they are often asymptomatic, but may present with cough, hemoptysis, or dyspnea. IMTs are low-grade tumors, histologically composedof inflammatoryspindle cells thatoftengrowlocallyandslowly [4]. A 42-year-old patient (para 0) presented after a positive homepregnancy test. Medical history was significant for an incidental lung lesion detected in 1989bychestX-ray followingapositivePPD.At that time, evaluation revealed a 2-cm, well-rounded, noncalcified lesion in the right lower lobe with no associated adenopathy. A biopsy (with preoperative evaluation showing negative hCG) was consistent with pseudotumor; no further treatment was advised. Several chest X-rays since 1989 showed a stable lesion. Obstetricgynecology history was noncontributory, with menarche at 13 years, regular periods, and no history of infertility. Upon presentation after positive pregnancy test, the patient reported only fatigue and shortness of breath upon exertion; she denied cough, sputum production, chest pain, fever, chills, or vaginal bleeding. Her last menstrual period was 51/2 weeks prior, and beta-hCG quantitative value (beta-quant) was 8961 mIU/mL. Multiple ultrasounds showed no intrauterine, molar, or ectopic pregnancy. Diagnostic laparoscopy confirmed no ectopic pregnancy; dilation and curettage revealed no gestational trophoblastic disease. The patient was given methotrexate; however, beta-quant continued to rise to 20 648mIU/mL over 5 days, and shewas referred togynecologic oncology.Gynecologic oncologyevaluated and referred her to thoracic surgery for assessment of her lung nodule. Lung exam was normal, and chest CT and positron emission tomography (PET) demonstrated a solitary, round, noncalcified right lower lobe nodule 1.2 cm (Fig. 1) with moderate uptake on PET. The patient underwent fiberoptic bronchoscopy and right thoracoscopic lower lobe wedge resection revealing a solitary, firm, solid lesion within the right lower lobe. Pathology revealed a noninvasive, 1-cm spindle cell lesion consistent with IMT. Postoperative beta-quant decreased steadily reaching zero approximately 3 weeks postoperatively, and the patient resumed regular periods. Beta-hCG secreting lung cancer is too rare to be a useful tumor marker by pulmonologists; however, it is critical that obstetriciangynecologists recognize the wide range of sources of beta-hCG production and refer patients for further evaluation after excluding common obstetric etiologies. The present case demonstrates the importance of considering all lung nodules as a source of ectopic betahCG; however, it is unclearwhether this case signifies a change in nature of the tumor, or a newfindingnot previously clinically evident. Given the recent evidence on multiple variants of hCG, further detailed analysis of hCG may be warranted in determining malignant potential of nontrophoblastic tumors. Physicians should be aware of the malignant nature ofmost beta-hCG secreting tumors and consider the potential for transformation of benign appearing nodules to more aggressive tumors.

Alteration of GABAergic synapses and gephyrin clusters in the thalamic reticular nucleus of GABAA receptor α3 subunit‐null mice

Multiple GABAA-receptor subtypes are assembled from alpha, beta and gamma subunit variants. GABAA receptors containing the alpha3 subunit represent a minor population with a restricted distribution in the CNS. In addition, they predominate in monoaminergic neurons and in the nucleus reticularis thalami (nRT), suggesting a role in the regulation of cortical function and sleep. Mice with a targeted deletion of the alpha3 subunit gene (alpha3(0/0)) are viable and exhibit a subtle behavioural phenotype possibly related to dopaminergic hyperfunction. Here, we investigated immunohistochemically the consequences of the loss of alpha3 subunit for maturation of GABAA receptors and formation of GABAergic synapses in the nRT. Throughout postnatal development, the regional distribution of the alpha1, alpha2, or alpha5 subunit was unaltered in alpha3(0/0) mice and the prominent alpha3 subunit staining of nRT neurons in wildtype mice was not replaced. Subcellularly, as seen by double immunofluorescence, the alpha3 and gamma2 subunit were clustered at postsynaptic sites in the nRT of adult wildtype mice along with the scaffolding protein gephyrin. In alpha3(0/0) mice, gamma2 subunit clustering was disrupted and gephyrin formed large aggregates localized at the cell surface, but unrelated to postsynaptic sites, indicating that nRT neurons lack postsynaptic GABAA receptors in mutant mice. Furthermore, GABAergic terminals were enlarged and reduced in number, suggesting a partial deficit of GABAergic synapses. Therefore, GABAA receptors are required for gephyrin clustering and long-term synapse maintenance. The absence of GABAA-mediated transmission in the nRT may have a significant impact on the function of the thalamo-cortical loop of alpha3(0/0) mice.

Luteinizing hormone, its beta-subunit variant, and epithelial ovarian cancer: the gonadotropin hypothesis revisited.

The gonadotropin hypothesis postulates that excessive gonadotropin stimulation results in increased proliferation and subsequent malignant transformation of ovarian epithelium. The authors evaluated this hypothesis by analyzing the association between serum levels of wild-type luteinizing hormone (LH) and ovarian cancer risk. They also examined the relation between a variant of LH containing two missense point mutations (Trp(8)Arg and Ile(15)Thr) in its beta-subunit and ovarian cancer risk. Fifty-eight cases of epithelial ovarian cancer and 116 controls matched on age, menopausal status, and date of blood donation were included in a case-control study nested within the New York University Women's Health Study, a prospective cohort enrolled between 1985 and 1991 in New York City. Wild-type serum levels and variant LH status were determined by immunofluorometric assays in which monoclonal antibodies specific for wild-type and variant LH were used. Compared with women in the lowest tertile of wild-type LH, women in the highest tertile had a lower risk of ovarian cancer, after adjustment for potential confounders (odds ratio = 0.42, 95% confidence interval: 0.09, 2.09). Women heterozygous for variant LH were not at increased risk (adjusted odds ratio = 0.95, 95% confidence interval: 0.27, 3.34). The results suggest that neither wild-type LH levels nor variant LH status is associated with increased risk of epithelial ovarian cancer.

Do antiepileptics phenytoin, carbamazepine, and loreclezole show GABA(A) receptor subtype selectivity in rat brain sections?

[35S]t-Butylbicyclophosphorothionate ([35S]TBPS), a convulsant site ligand of GABA(A) receptors, was used in autoradiography with rat brain sections to test suggested receptor subtype-selective actions of antiepileptics phenytoin, carbamazepine and loreclezole on native GABA(A) receptors. At maximal 100 microM concentration, both phenytoin and carbamazepine decreased [35S]TBPS binding only by 20%, indicating that their low potency and efficacy prevents their use as alpha1 subunit-identifying compounds. Ten microM loreclezole did not affect the binding, but a further increase in loreclezole concentration strongly decreased it. The action of loreclezole, assumed to reflect beta2/3 subunit-containing receptors, varied from brain region to region, but the effects were unrelated to the regional expression profiles of beta subunit variants. We conclude that in autoradiographic [35S]TBPS binding assay neither carbamazepine, phenytoin nor loreclezole are useful tools in characterizing brain regional heterogeneity of GABA(A) receptors in rats and that only loreclezole exhibits high, pharmacologically relevant efficacy.

Expression of Ca(2+) channel subunits during cardiac ontogeny in mice and rats: identification of fetal alpha(1C) and beta subunit isoforms.

Functional cardiac L-type calcium channels are composed of the pore-forming alpha(1C) subunit and the regulatory beta(2) and alpha(2)/delta subunits. To investigate possible developmental changes in calcium channel composition, we examined the temporal expression pattern of alpha(1C) and beta(2) subunits during cardiac ontogeny in mice and rats, using sequence-specific antibodies. Fetal and neonatal hearts showed two size forms of alpha(1C) with 250 and 220 kDa. Quantitative immunoblotting revealed that the rat cardiac 250-kDa alpha(1C) subunit increased about 10-fold from fetal days 12-20 and declined during postnatal maturation, while the 220-kDa alpha(1C) decreased to undetectable levels. The expression profile of the 85-kDa beta(2) subunit was completely different: beta(2) was not detected at fetal day 12, rose in the neonatal stage, and persisted during maturation. Additional beta(2)-stained bands of 100 and 90 kDa were detected in fetal and newborn hearts, suggesting the transient expression of beta(2) subunit variants. Furthermore, two fetal proteins with beta(4) immunoreactivity were identified in rat hearts that declined during prenatal development. In the fetal rat heart, beta(4) gene expression was confirmed by RT-PCR. Cardiac and brain beta(4) mRNA shared the 3 prime region, predicting identical primary sequences between amino acid residues 62-519, diverging however, at the 5 prime portion. The data indicate differential developmental changes in the expression of Ca(2+) channel subunits and suggest a role of fetal alpha(1C) and beta isoforms in the assembly of Ca(2+) channels in immature cardiomyocytes.

Differential regulation of synaptic GABAA receptors by cAMP-dependent protein kinase in mouse cerebellar and olfactory bulb neurones.

1. It has been demonstrated that the regulation of recombinant GABAA receptors by phosphorylation depends on the subunit composition. Here we studied the regulation of synaptic GABAA receptor function by cAMP-dependent protein kinase (PKA) in neurones expressing distinct receptor subtypes. 2. Light microscopic immunocytochemistry revealed that granule cells of the olfactory bulb express only the beta3 as the beta subunit variant, whereas cerebellar stellate and basket cells express only the beta2 as the beta subunit. 3. In cerebellar interneurones, intracellular application of 20 microM microcystin, a protein phosphatase 1/2A inhibitor, prolonged (63 +/- 14 %; mean +/- s.e.m.) the decay time course of miniature IPSCs (mIPSCs) without significantly affecting their amplitude, rise time and frequency. The effect of microcystin could be blocked by co-applying PKA inhibitory peptide (PKA-I, 1 microM). 4. No significant changes in any of the mIPSC parameters could be detected after intracellular application of PKA-I alone or following the inhibition of calcineurin with FK506 (50 nM). 5. In granule cells of the olfactory bulb expressing the beta3 subunit fast and slowly rising mIPSCs were detected, resulting in a bimodal distribution of the 10-90 % rise times, suggesting two distinct populations of events. Fast rising mIPSCs (mIPSCFR) had a 10-90 % rise time of 410 +/- 50 micros, an amplitude of 68 +/- 6 pA, and a weighted decay time constant (tauw) of 15.8 +/- 2.9 ms. In contrast, slowly rising mIPSCs (mIPSCSR) displayed an approximately threefold slower rise time (1.15 +/- 0.12 ms), 57 % smaller amplitude (29 +/- 1.7 pA), but had a tauw (16.8 +/- 3.0 ms) similar to that of the fast events. 6. mIPSCs in olfactory granule cells were not affected by the intracellular perfusion of microcystin. In spite of this, intracellular administration of constitutively active PKA caused a small, gradual, but significant increase (18 +/- 5 %) in the amplitude of the events without changing their time course. 7. These findings demonstrate a cell-type-dependent regulation of synaptic inhibition by protein phosphorylation. Furthermore, our results show that the effect of PKA-mediated phosphorylation on synaptic inhibition depends upon the subunit composition of postsynaptic GABAA receptors.

The beta enolase subunit displays three different patterns of microheterogeneity in human striated muscle.

In higher vertebrates, the glycolytic enzyme enolase (2-phospho-D-glycerate hydrolyase; EC 4.2.1.11) is active as a dimeric protein formed from three subunits--alpha: ubiquitous, beta: muscle specific, and gamma: neuron specific--encoded by different genes. In the present study, we have shown that an antiserum previously produced against the mouse beta beta enolase is also a specific reagent for the muscle specific human enolase. Using this antiserum to study human muscles, we demonstrated novel patterns of the beta subunit microheterogeneity which are distinctive from those observed previously in rodents and which appear to be independent of age, gender and muscular activity. Two variants of the beta subunit differing by their size have been detected: one heavy form of 46 kDa (beta H) and one light form of 45 kDa (beta L). Muscle biopsies expressed either beta H or beta L or beta H + beta L, and all muscles of an individual expressed the same variants. The products of in vitro translation of RNA prepared from human muscle displayed beta subunit variants identical to those of the protein present in the biopsy. Therefore the differences observed between individuals reveal a difference already present at the level of the RNA transcripts. These observations suggest the existence of an yet undescribed polymorphism of the human beta enolase gene which could affect the coding sequence. Comparative immunocytochemical and histochemical analyses of biopsies demonstrated that the beta subunit was expressed in all fast fibres (type II), but not in slow fibres (type I). No difference was observed in the intensity of beta enolase immunolabelling between the various types (IIA, IIAB, IIB) of fast fibres. No significant difference in fibre type composition and histological appearance was visible between muscles presenting either one of the three patterns of microheterogeneity.

Increased prevalence of luteinizing hormone beta-subunit variant in Japanese infertility patients.

To determine the frequency of a common luteinizing hormone variant in a Japanese population and to evaluate its significance in infertility, serum samples were collected from 169 healthy non-pregnant Japanese women, 105 healthy adult Japanese men and 97 female Japanese infertility patients. The luteinizing hormone variant includes two point mutations in the beta-subunit gene (Trp8 to Arg8 and Ile15 to Thr15). DNA from blood cells was studied in 10 healthy women, 10 men and five patients using polymerase chain reaction and direct sequencing. In immunoassays, results with a monoclonal antibody recognizing only the wild-type hormone and a polyclonal antibody recognizing the variant as well were compared as a ratio; ratios in heterozygotes and in individuals with only wild-type alleles ranged from 0.19 to 0.50 and from 0.56 to 1.21, respectively, and 0.50 was considered a 'cut-off' value for identifying individuals with the variant. For the larger subject groups, the frequency of the variant was 9.5% in normals. The mean ratio (0.80 +/- 0.35) in infertility patients was significantly lower (P < 0.01) than in healthy women (1.09 +/- 0.56), and the variant occurred more frequently in infertility patients (16.5%) than in healthy women (8.3%; P < 0.05). The variant was more frequent in patients with ovulatory disorders (43.8%) than other patients (16.0%; P < 0.05).

Subtype specificity of gamma-aminobutyric acid type A receptor antagonism by clozapine.

Clozapine, an atypical neuroleptic, functionally antagonizes the gamma-aminobutyric acid-induced chloride uptake via the main central inhibitory receptor, gamma-aminobutyric acid type A (GABAA) receptor, in brain vesicles. GABAA antagonism by micromolar concentrations of clozapine is more efficient in rat cerebrocortical and hippocampal membranes than in cerebellar membranes, as evidenced by clozapine reversal GABA-inhibition of [35S]t-butylbicyclophosphorothionate ([35S]TBPS) binding. A typical neuroleptic, haloperidol, failed to antagonize GABA in any of these brain regions, while the specific GABAA antagonist 2'-(3'-carboxy-2',3'-propyl)-3-amino-6-p -methoxyphenylpyrazinium bromide (SR 95531) was efficient in all three brain regions. Clozapine action on [35S]TBPS binding was unaffected by the benzodiazepine receptor antagonist flumazenil. Clozapine inhibited the binding of [3H]muscimol and [3H]SR 95531 to the GABA recognition site, but this effect only partially correlated with the regional differences in and the potency of clozapine antagonism of GABA-inhibition of [35S]TBPS binding, suggesting that also other than GABA sites may mediate clozapine actions. Autoradiography of [35S]TBPS binding revealed GABA antagonism by clozapine in most brain regions. Main exceptions were cerebellar granule cell and molecular layers, olfactory bulb external plexiform and glomerular layers and primary olfactory cortex, where clozapine antagonized GABA inhibition less than average, and lateral hypothalamic and preoptic areas where its antagonism was greater than average. Recombinant alpha 6 beta 2 gamma 2 receptors, the predominant alpha 6 subunit-containing receptor subtype in cerebellar granule cells, failed to show GABA antagonism by clozapine up to 100 microM. In contrast, recombinant alpha 1 beta 2 gamma 2 receptors, forming the predominant receptor subtype in the brain, were clozapine sensitive. Recombinant alpha 6 beta 2 gamma 2 and alpha 6 beta 3 gamma 2 receptors resulted in clozapine-insensitive receptors, whereas alpha 6 beta 1 gamma 2 receptors were clozapine sensitive. The efficacy of clozapine to antagonize GABA in alpha 1 beta x gamma 2 receptors decreased in the order of alpha 1 beta 1 gamma 2 > alpha 1 beta 2 gamma 2> alpha 1 beta 3 gamma 2. The results indicate that clozapine antagonizes the function of most GABAA receptor subtypes, and that the interaction is determined by the interaction of the alpha and beta subunit variants. GABA antagonism is a unique property of clozapine, not shared by haloperidol, which might be involved in the pharmacological mechanism for the increased seizure susceptibility associated with clozapine treatment.

Prevalence of an immunological LH β‐subunit variant in a UK population of healthy women and women with polycystic ovary syndrome

An immunological LH beta-subunit variant has been described, which is undetectable using monoclonal antibodies directed to the intact LH molecule alone. Subjects have been found homozygous or heterozygous for nucleotide mutations within codons 8 and 15 in the LH beta-subunit gene. The prevalence of the variant LH beta-subunit has been estimated in a healthy UK population of women of reproductive age and in women with polycystic ovary syndrome (PCOS). The relationship of the variant molecule to the clinical and hormonal parameters of the subjects has been evaluated. The control and PCOS subjects were screened for the presence of the mutation by using a ratio of two immunofluorometric assays using monoclonal antibodies (Mab). One assay, not detecting the LH variant, uses a Mab directed to the intact LH molecule and a beta-specific Mab. The other assay, detecting both the variant and wild-type LH, uses two beta-subunit specific Mabs. The mutations in the LH beta-subunit gene were confirmed by restriction fragment length polymorphism. The relationship of the presence of the variant to the clinical and hormonal parameters was assessed by ANOVA. Two hundred and twelve normal ovulatory women, of whom 66 (31%) were obese (body mass index > 25) and 146 (69%) non-obese, and 153 women with PCOS, 115 (75%) obese and 38 (25%) non-obese participated in the study. The variant LH was detected in 31 (15%) controls and 32 (21%) PCOS subjects (P = 0.124) using specific Mab. Obese PCOS had a higher incidence of the heterozygous LH variant compared to obese controls (odds ratio 2.5, P = 0.03), and compared to non-obese PCOS (odds ratio 6.3, P = 0.01). The previously described two mutations in codon 8 and codon 15 were present in all subjects detected to be mutant hetero of homo-zygous by RFLP. There was no relationship between the presence of the variant LH and the clinical and hormonal parameter in the PCOS subjects; however, in the controls the presence of the variant LH was associated with a higher serum total testosterone (P = 0.046), oestradiol (P = 0.03) and SHBG (P = 0.002). The results of this study show that the variant LH beta-subunit is a common polymorphism occurring in 15% of a healthy UK population of women. The prevalence was not higher in women with PCOS, though it was over represented in obese women with PCOS. The presence of the variant did not alter the clinical or hormonal expression of the disorder in women with PCOS. Its presence in the controls was however associated with higher serum oestradiol and probably secondary elevation of SHBG and testosterone, suggesting that the variant form of LH may be associated with subtle changes in the function of the hypothalamic-pituitary-gonadal axis.

Insulin-like growth factor I receptors of fetal brain are enriched in nerve growth cones and contain a beta-subunit variant.

Nerve growth cones isolated from fetal rat brain are highly enriched in a 97-kDa glycoprotein, termed beta gc, that comigrates with the beta subunit of the IGF-I receptor upon two-dimensional PAGE and is disulfide-linked to this receptor's alpha subunit. Antibodies prepared to a conserved domain shared by the insulin and IGF-I receptor beta subunits (AbP2) or to beta gc were used to study receptor distribution further. Subcellular fractionation of the fetal brain segregated most AbP2 immunoreactivity away from growth cones, whereas most beta gc immunoreactivity copurified with growth cones. Experiments involving ligand-activated receptor autophosphorylation confirmed the concentration of IGF-I but not of insulin receptors in growth cone fractions. These results indicate the enrichment of IGF-I receptors in (presumably axonal) growth cones of the differentiating neuron. Furthermore, the segregation of beta gc from AbP2 immunoreactivity suggests that such neurons express an immunochemically distinct variant of the IGF-I receptor beta subunit at the growth cone.

Distribution, prevalence, and drug binding profile of gamma-aminobutyric acid type A receptor subtypes differing in the beta-subunit variant.

Native gamma-aminobutyric acid type A (GABAA) receptors containing different beta-subunit variants were identified immunobiochemically with antisera recognizing selectively the beta 1-, beta 2-, or beta 3-subunit. As determined by immunoprecipitation, the beta 2-subunit was present in 55-60% of GABAA receptors, while only minor receptor populations contained the beta 1-subunit (16-18%) or the beta 3-subunit (19-25%). Since the sum of these values amounts to about 100%, it is concluded that GABAA receptors largely contain only a single type of beta-subunit. Pharmacologically, receptors containing the beta 2-subunit differed from those containing the beta 1- or beta 3-subunit by their differential affinities for benzodiazepine receptor ligands. The subunit composition was analyzed biochemically in receptors immunoprecipitated by the beta 2-subunit antiserum. The beta 2-subunit was preferentially associated with the alpha 1-subunit (rarely with the alpha 2-subunit) and with the gamma 2-subunit; negligible or no immunoreactivity was detected for the alpha 3-, alpha 5-, or beta 1-subunit. A stringent co-expression of alpha 1- and beta 2-subunits was confirmed by double immunofluorescence staining on the cellular level. Neurons expressing the beta 3-subunit immunoreactivity were largely double labeled by the alpha 2-subunit antiserum. Thus, the subunit combinations alpha 1 beta 2 gamma 2 and alpha 2 beta 3 gamma 2 represent two main GABAA receptor subtypes, which together amount to 75-85% of the diazepam-sensitive GABAA receptors.

A single histidine in GABAA receptors is essential for benzodiazepine agonist binding.

Benzodiazepines (BZ) modulate neurotransmitter-evoked chloride currents at the gamma-aminobutyric acid type A (GABAA) receptor, the major inhibitory ion channel in the mammalian brain. This receptor is composed of structurally distinct subunits whose numerous molecular variants underlie the observed diversity in the properties of the BZ site. Pharmacologically distinct BZ sites can be recreated by the recombinant coexpression of any one of six alpha subunits, a beta subunit variant, and the gamma 2 subunit. In these receptors the alpha variant determines the affinity for ligand binding of the BZ site. Notably, the alpha 1 and alpha 6 variants impart on alpha chi beta 2 gamma 2 receptors high and negligible affinity, respectively, to BZ ligands with sedative as well as anxiolytic activities. By exchanging domains between the alpha 1 and alpha 6 variants, we show that a portion of the large extracellular domain determines sensitivity toward these ligands. Furthermore, we identify a single histidine residue in the alpha 1 variant, replaced by an arginine in alpha 6, as a major determinant for high affinity binding of BZ agonists. This residue also plays a role in determining high affinity binding for BZ antagonists. Hence, this histidine present in the alpha 1, alpha 2, alpha 3, and alpha 5 subunits appears to be a key residue for the action of clinically used BZ ligands.

Association of Hemoglobin H Disease With Hb J-Iran (β77 His → Asp) Impact on Subunit Assembly

A young Iranian female was found to be heterozygous for hemoglobin (Hb) J-Iran (beta 77 His----Asp) in combination with Hb H disease. The proportion of Hb J in the patient's hemolysate was surprisingly high: 65% Hb J, 30% Hb A. Thus, the interaction of a negatively charged beta subunit variant of Hb with alpha-thalassemia leads to a marked increase in the relative amount of the variant Hb within red cells. This observation provides further support for an electrostatic model of Hb subunit assembly.

Association of hemoglobin H disease with Hb J-Iran (beta 77 His---- Asp): impact on subunit assembly

A young Iranian female was found to be heterozygous for hemoglobin (Hb) J-Iran (beta 77 His----Asp) in combination with Hb H disease. The proportion of Hb J in the patient's hemolysate was surprisingly high: 65% Hb J, 30% Hb A. Thus, the interaction of a negatively charged beta subunit variant of Hb with alpha-thalassemia leads to a marked increase in the relative amount of the variant Hb within red cells. This observation provides further support for an electrostatic model of Hb subunit assembly.