Biomedical subjects

F C Chang

Publications and source records attributed to F C Chang.

At least 37 records · Page 2Linked to original sources

PubMed2000

"Paper-clip" type triple helix formation by 5'-d-(TC)3Ta(CT)3Cb(AG)3 (a and b = 0-4) as a function of loop size with and without the pseudoisocytosine base in the Hoogsteen strand.

The formation of a DNA "paper-clip" type triple helix (triplex) with a common sequence 5'-d-(TC)(3)T(a)()(CT)(3)C(b)()(AG)(3) (a and b = 0-4) was studied by UV thermal melting experiments and CD spectra. These DNA oligomers form triplexes and duplexes under slightly acidic and neutral conditions, respectively. The stability of the formed triplexes (at pH 4.5) or duplexes (at pH 7.0 or 8.0) does not vary significantly with the size of the loops (a and b = 1-4). At pH 6.0, the triplex stability is, however, a function of a and b. It is also interesting to note that the oligomer 5'-d-(TC)(3)(CT)(3)(AG)(3) (a and b = 0) forms a stable triplex at pH 4.5 with a slightly lower T(m) value, due to dissociation of a base triad at one end and a distorted base triad at the other, observed by (1)H NMR. Thus, we have here a model system, 5'-d-(TC)(3)T(a)(CT)(3)C(b)(AG)(3), that could form a triplex effectively with (a and b = 1-4) and without (a and b = 0) loops under acidic conditions. In addition, the triplex formation of oligomers with replacement of one, two, or three 2'-deoxycytidine in the Hoogsteen strand by either 2'-deoxypseudoisocytidine (D) or 2'-O-methylpseudoisocytidine (M) was also studied in the sequence 5'-d-(TX)(3)T(2)(CT)(3)C(2)(AG)(3) (where X is C, D, or M). Both CD spectra and UV melting results showed that only D3 [(TX)(3) = (TD)(3)] and M3 [(TX)(3) = (TM)(3)] were able to form the paper-clip structure under both neutral and acidic conditions. This is because the N(3)H of a pseudoisocytosine base can serve as a proton donor without protonation. We hereby proved that the 2'-deoxypseudoisocytidine, similar to 2'-O-methylpseudoisocytidine, could replace 2'-deoxycytidine in the Hoogsteen strand to provide triplex formation at neutral pH.

Adenine↗

PubMed2000

The identification of an endonuclease that cleaves within an HuR binding site in mRNA.

Messenger RNAs (mRNAs) that contain U-rich elements are targeted for rapid decay. Selective inhibition of this decay results in a rapid increase in steady state level. Thus, this is an important regulatory step in gene expression. Previously, we have found that these mRNAs are selectively stabilized by a specific mRNA binding protein called HuR. The mechanism of action of HuR is not well understood. It has been postulated that HuR stabilizes mRNA by the displacement or inhibition of factors that specifically cleave or deadenyl-ate these mRNAs. In this paper, we report the identification and characterization of a novel endo-nuclease that cleaves within an HuR binding site in p27kip1 mRNA. The specificity of this endonuclease and its inhibition by HuR argue for it playing a role in the postranscriptional regulation of gene expression.

Antigens, Surface↗

PubMed2000

Debt and other influences on career choices among surgical and primary care residents in a community-based hospital system.

BACKGROUND: To evaluate debt and other factors that help formulate the career paths of future surgical and primary care physicians, a survey was undertaken. METHODS: Surgical specialty (SS) and primary care (PC) residents were surveyed regarding demographics, factors influencing choice of specialty, methods of financing education, debt characteristics, and outlooks regarding future earnings and practice characteristics. RESULTS: The clinical years of medical school and personalities of specialists and residents were important factors in career choices for both PC and SS. The length of residency, desirable lifestyle, and working hours were all more important to PC residents. Surgeons found intellectual challenge and procedure-based practice of greater importance. Although not highly regarded by either group, scholarship obligation and student loans had a significantly greater impact on specialty choice and practice plans for PC residents. At the completion of training, 55% of SS and 28% of PC residents anticipate owing more than $100,000. Debt was especially significant in specialty choice and practice plans for PC residents with debt over $100,000. CONCLUSION: Surgical residents are less concerned about personal sacrifices in their quest to become surgeons. It appears state funded scholarships are successful in attracting students to primary care. Both SS and PC residents have significant debt, although, SS residents have greater financial debt than primary care residents. However, the anticipation of indebtedness was a more significant factor in determination of career path for PC.

Adult↗

PubMed2000

Routine blastocyst culture and transfer: 201 patients' experience.

PURPOSE: The purpose was to compare the outcomes between day-5 blastocyst and day-2 embryo transfers. METHODS: Infertile women who accepted the ovarian hyperstimulation and oocyte retrieval were divided: Group 1, day 2 embryo transfer, group 2, cultured to day 5 in serum-free sequential culture medium and transfer. Early embryo quality and growth, blastocyst formation and quality, implantation rate (IR) and pregnancy rates (PR) were detected. RESULTS: Total blastocyst formation rate was 49.4%. Better early embryo quality (days 2, 3) and higher day 3 blastomere number possess higher blastocyst formation rate. The IR for day 2 and day 5 embryos were 10.8% and 22.2%, respectively. The PR in both groups were comparable (37.3% vs. 41.8%). CONCLUSIONS: Blastocyst transfer has higher IR and comparable PR as those of day 2 embryo transfer. Early embryo qualities and day 3 blastomere number are useful in predicting the final blastocyst formation. Blastocyst formation rate is not related to maternal age, infertile causes, insemination methods, and early embryo number.

Adult↗

PubMed2000

Longitudinal survey of fetal ventricular ejection and shortening fraction throughout pregnancy.

OBJECTIVES: To assess and compare the ejection fraction (EF) and shortening fraction (SF) of both ventricles through pregnancy. MATERIALS: Uncomplicated singleton pregnancies, of between 10 and 40 weeks' gestation, were recruited. Transabdominal two-dimensional M-mode imaging of the right ventricular EF (REF), right ventricular SF (RSF), left ventricular EF (LEF), and left ventricular SF (LSF) were performed. Doppler surveys were performed at (1) 10-14 weeks; (2) 15-19 weeks; (3) 20-24 weeks; (4) 25-29 weeks; (5) 30-34 weeks; and (6) 35-40 weeks. The following values were measured: REF, RSF, LEF, LSF, REF/LEF and RSF/LSF. RESULTS: A total of 42 cases were included. The values of REF, RSF, LEF, and LSF for the six gestational time periods were (1) 0.86, 0.49, 0.85, 0.48; (2) 0.84, 0.48, 0.88, 0.51; (3) 0.76, 0.37, 0.84, 0.48; (4) 0.75, 0.37, 0.84, 0.48; (5) 0.76, 0.38, 0.83, 0.47; and (6) 0.77, 0.38, 0.84, 0.48, respectively. The ratios of REF/LEF and RSF/LSF in each gestational time period were: (1) 1.04, 1.11; (2) 0.95, 0.92; (3) 0.95, 0.90; (4) 0.94, 0.90; (5) 0.94, 0.92; and (6) 0.92, 0.89, respectively. CONCLUSIONS: After 20 weeks' gestation both the EF and SF of the right ventricle decreased. A non-significant change in the LEF and LSF was observed throughout pregnancy. A decreasing trend in the REF/LEF and RSF/LSF ratios during pregnancy was noted.

Adult↗

PubMed2000

Doppler evaluation of the uterine and spiral arteries from different sampling sites and phases of the menstrual cycle during controlled ovarian hyperstimulation.

OBJECTIVE: To compare the pulsatility index (PI) and resistance index (RI) at different sampling sites of the uterine and spiral arteries in the early and mid-menstrual phases. MATERIALS AND METHODS: The uterine and spiral arteries of 110 women undergoing similar ovarian hyperstimulation and intra-uterine insemination regimes were examined using transvaginal color Doppler ultrasound. The uterine artery was sampled at five sites: (1) ascending branch; (2) descending branch; (3) proximal, near branch division; (4) mid, 0.5 cm distal to the division; (5) lateral location, 1 cm distal to the division. The spiral artery was sampled at three sites: (a) anterior; (b) fundal; (c) posterior. The uterine and spiral arteries were examined twice, on days 2-3 and 14-16, respectively, of the menstrual cycle. The women were also grouped according to age at examination, < or = 30 years and > 30 years. The PI and RI values for different sites, menstrual phase, and age were compared. RESULTS: The mean PI and RI values of the uterine artery were: (1) 2.86 +/- 1.20 and 0.92 +/- 0.13; (2) 2.66 +/- 1.15 and 0.89 +/- 0.12; (3) 2.88 +/- 1.26 and 0.90 +/- 0.15; (4) 3.03 +/- 1.02 and 0.91 +/- 0.07; (5) 3.23 +/- 1.38 and 0.89 +/- 0.12; and of the spiral artery were (a) 1.61 +/- 1.01 and 0.69 +/- 0.17; (b) 1.69 +/- 0.74 and 0.74 +/- 0.17; (c) 1.73 +/- 0.86 and 0.68 +/- 0.17. The PI values for uterine and spiral arteries at two phases of the menstrual cycle were 2.92 +/- 1.18 and 1.55 +/- 0.72 (days 2-3); 3.11 +/- 1.15 and 1.80 +/- 1.02 (days 14-16), respectively; for younger women (age < or = 30 years) these values were 2.83 +/- 1.22 and 1.6 +/- 0.85 and for older women (age > 30 years) 3.0 +/- 1.34 and 1.72 +/- 0.96, respectively. CONCLUSIONS: There were no significant differences in PI and RI values of the uterine and spiral arteries at different sampling sites, phase of the menstrual cycle or age. The higher PI values tended to occur in the lateral uterine artery and posterior spiral artery, during the mid-menstrual phase and in the older age group. The PI and RI values of the mid-uterine and fundal spiral artery sampling sites are representative of the whole uterine artery and spiral artery, respectively.

Adult↗

PubMed2000

Salvage surgery for recurrent nasopharyngeal carcinoma.

OBJECTIVE: To evaluate the efficacy of salvage surgery in the treatment of recurrent nasopharyngeal carcinoma (NPC) at the primary site. STUDY DESIGN: A retrospective investigation of the outcome of salvage surgery for 28 patients with recurrent NPC after definite radiation therapy. METHODS: The nasopharynx was approached anteroposteriorly by the transmaxillary approach (maxillary swing, maxillectomy) or inferior approach (midline mandibulotomy or median labiomandibular glossotomy), or laterally by modified facial translocation or transpterygoid approach; intentional ligation of the internal carotid artery was performed after establishment of extracranial-intracranial (EC-IC) bypass in one patient; postoperative irradiation was given to the patients with positive pathological margins. RESULTS: Nine patients lived without disease for 20 to 93 months (mean interval, 52 mo) after surgery; among them, eight patients had T1 tumors that were resected totally by surgery via anteroposterior approaches and the other patient had postoperative irradiation to control the disease. Seven patients had local recurrence 8 to 21 months after treatment. Four patients developed distant metastases, including one patient with a T2b tumor that was totally resected through modified facial translocation approach with ligation of internal carotid artery. Eight patients died of other causes; internal carotid artery blowout was the cause of death in four of these eight patients. CONCLUSIONS: In most cases of recurrence, T1 nasopharyngeal tumors can be resected totally by anteroposterior approaches; for T2 or larger tumors, postoperative irradiation is usually necessary. Otherwise, facial translocation offers a better chance to completely resect the tumors. Internal carotid artery is better ligated if patients have received greater than 70 Gy irradiation or if the artery must be exposed during the surgery. We suggest that EC-IC bypass be used to avoid the possible complications (or cerebral ischemic stroke) caused by ligation of internal carotid artery. The transmaxillary approach is favored in the management of nasopharyngeal tumor recurrence with nasal cavity extension, and midline mandibulotomy is more suitable for resection of posterior margin of nasopharyngeal tumor recurrence. Facial translocation offers the widest operative field and is the most versatile approach for radical resection of nasopharyngeal tumor recurrence, but the surgeon should be skilled in the management of the facial nerves to reduce morbidity.

Adult↗

PubMed2000

IL-1 is a mediator of increases in slow-wave sleep induced by CRH receptor blockade.

We hypothesize that corticotropin-releasing hormone (CRH), a regulator of the hypothalamic-pituitary-adrenal (HPA) axis, is involved in sleep-wake regulation on the basis of observations that the CRH receptor antagonist astressin, after a delay of several hours, reduces waking and increases slow-wave sleep (SWS) in rats. This delay suggests a cascade of events that begins with the HPA axis and culminates with actions on sleep regulatory systems in the central nervous system. One candidate mediator in the brain for these actions is interleukin (IL)-1. IL-1 promotes sleep, and glucocorticoids inhibit IL-1 synthesis. In this study, central administration of 12.5 microgram astressin into rats before dark onset reduced corticosterone 4 h after injection and increased mRNA expression for IL-1alpha and IL-1beta but not for IL-6 or tumor necrosis factor-alpha in the brain 6 h after injection. To determine directly whether IL-1 is involved in astressin-induced alterations in sleep-wake behavior, we then pretreated rats with 20 microgram anti-IL-1beta antibodies before injecting astressin. The increase in SWS and the reduction in waking that occur after astressin are abolished when animals are pretreated with anti-IL-1beta. These data indicate that IL-1 is a mediator of astressin-induced alterations in sleep-wake behavior.

Animals↗

PubMed2000

Effect of oxidized frying oil and vitamin C levels on the hepatic xenobiotic-metabolizing enzyme system of guinea pigs.

The influence of oxidized frying oil (OFO) on the guinea pig hepatic microsomal xenobiotic-metabolizing enzyme system in the presence of different amounts dietary vitamin C was investigated. Weanling male guinea pigs were divided into four groups and were fed 15% oxidized frying oil diets supplemented with vitamin C at 300, 600, or 1,500 mg/kg (experimental diets) or a control diet that contained 15% fresh untreated soybean oil with 300 mg/kg of vitamin C, respectively. After 60 d, guinea pigs were euthanized and phase I and phase II xenobiotic-metabolizing enzymes in the liver were determined. Compared with the fresh oil diet fed the control group, the relative liver weight was higher in the OFO-fed groups. Hepatic microsomal protein and cytochrome P450 contents were significantly higher in OFO-fed guinea pigs than in the control group. Both values increased in response to increased intake of vitamin C. The activities of phase II relative components, including UDP-glucuronyl transferase, UDP-glucuronyl dehydrogenase and beta-glucuronidase, of guinea pigs fed the OFO diets supplemented with 300 mg vitamin C/kg were significantly higher than those of guinea pigs fed the control diet. However, the phase II relative components decreased with increasing vitamin C content in the diet. The results demonstrate that both dietary OFO and vitamin C in guinea pigs induce hepatic xenobiotic-metabolizing enzymes, but the level of induction is modulated by the dietary vitamin C level.

Animals↗

PubMed2000

Imaging of invasiveness of pituitary adenomas.

The purpose of our study was to examine the tumor size, imaging invasiveness of the pituitary macroadenomas (PMA) and to evaluate the directions of PMA spread. One hundred and thirty-five patients with PMA were examined with MRI and/or CT for pre-operative evaluation. We retrospectively reviewed the CT and MRI to identify tumor size, extension and to evaluate the directions of tumor spread. One hundred and seventeen patients (87%) had suprasellar extension with compression of optic apparatuses, twelve patients (9%) had extension of tumor upward to hypothalamus and third ventricle. Infrasellar extension via the floor of the sella and sphenoid sinus was found in thirty-eight patients (28%), and further downward extension to ethmoid sinus, nasopharynx and/or skull base was depicted in five patients (4%). Twenty-two patients (16%) had lateral invasion to the cavernous sinus and associated cranial nerves. Temporal and frontal extensions were depicted in seven patients (5%) and six patients (4%), respectively. Five patients (4%) had posterior subtentorial extension to posterior fossa. Histologically, only two patients showed microscopic invasive features. There was no correlation between histologic features and imaging invasiveness. The PMA had the potential of multi-directional extension. This experience indicated any type of pituitary adenoma could invade surrounding structures. Suprasellar invasion was the most common direction of pituitary adenoma spread, followed by infrasellar, lateral, anterior and posterior routes.

Adenoma↗

PubMed2000

Endovascular embolization of intractable epistaxis.

BACKGROUND: We undertook this study to define the role of angiography and endovascular embolization in the treatment of patients with intractable epistaxis. METHODS: A series of 19 patients with massive intractable epistaxis, all treated with endovascular embolization were reviewed. There were 15 males and four females ranging in age from 18 to 70 years, with a mean age of 38 years. The predisposing factors of intractable epistaxis were head and neck tumors (n = 11), idiopathic symptoms (n = 3), surgical complications (n = 2), arteriovenous malformation of the face (n = 1), thrombocytopenia (n = 1) and trauma (n = 1). Polyvinyl alcohol (PVA) particles and gelfoam plugs were used as embolic agents in 11 patients for devascularization; detachable balloons with N-butyl-2-cyanoacrylate (NBCA) were used in four patients for vascular occlusion; NBCA and/or coils were used in four patients in the carotid and subclavian arteries for obliteration of pseudoaneurysms. RESULTS: Complete cessation of epistaxis was achieved in all 19 patients immediately after embolization. Seven patients had also undergone surgical tumor removal at two to five days after embolization. Two patients experienced mild to moderate facial pain on the first day after the procedure, but the symptoms later subsided. No significant complication or recurrence was observed in 18 patients. One patient with advanced hypopharyngeal carcinoma died two weeks after embolization due to another episode of massive epistaxis. Clinical follow-up for these patients was 15 days to eight years, with a mean of 41 months. CONCLUSIONS: Therapeutic endovascular embolization of intractable epistaxis is both efficient and safe. It should be considered as the primary treatment modality in intractable epistaxis.

Adolescent↗

PubMed2000

Gender prevalence in twin-twin transfusion syndrome.

BACKGROUND: This study aimed to determine the gender prevalence of fetuses complicated with twin-twin transfusion syndrome (TTTS). METHODS: All cases of TTTS corresponded with the following major criteria: a single placenta, monochorion, the same gender, and a combination of polyhydramnios-oligohydramnios. At least one of three minor criteria were required for the establishment of TTTS, including a stuck twin, a birth weight discordance exceeding 20%, and hemoglobin difference > 5 g/dl. RESULTS: Fifty-six twin pregnancies met the above criteria, of which 33 (58.9%) twin pairs were female. The female tendency existed, but there was a non-significant difference. Mean gestational age at diagnosis was 20.2 +/- 3.2 weeks. The birth weight discordance exceeding 20% was present in 50 of 56 (89.3%), and mean growth discordance was 32% +/- 8%. A stuck twin was noted in 37 of 56 cases (66.1%). The mortality of fetuses or neonates was 34.8% (39/112), including 8 (7.1%) fetal deaths and 31 (27.6%) neonatal deaths. There were no differences in maternal age, parity, or gestational age of delivery between male and female pregnancies. CONCLUSION: Although the female preponderance did not reach statistical significance, the female tendency might still exist after a larger series analysis. The female tendency may be the result of the gender difference in monochorionic twins. The gender difference could provide research implications and a diagnostic warning for clinicians in monochorionic twin pregnancies before the presence of TTTS.

Female↗

PubMed2000

Double-balloon technique for embolization of carotid cavernous fistulas.

Embolization of a carotid cavernous fistula (CCF) by means of a detachable balloon is an established method for treating CCFs while preserving a patent parent internal carotid artery (ICA). However, failure to embolize the CCF may occur on a few occasions, such as when the balloon cannot pass through the fistula into the cavernous sinus by blood flow, or when the inflated balloon in the cavernous sinus retracts to the carotid artery. Under these circumstances, the ICA may have to be sacrificed in order to treat the CCF. Herein we describe a double-balloon technique for embolization of a CCF. By applying this technique, we successfully treated nine of 11 CCFs, without compromise of the parent ICA when the conventional one-balloon technique failed.

Balloon Occlusion↗

PubMed2000

Accuracy of sonography in predicting the outcome of fetal congenital diaphragmatic hernia.

BACKGROUND: The outcome of congenital diaphragmatic hernia (CDH) remains poor despite recent advances in neonatal care. This study was designed to evaluate the role of sonography in predicting the outcome of CDH. METHODS: Pregnancies with CDH were studied. Fetal survival, morbidity, combined anomalies and mortality were recorded. Seven parameters were recorded, including the presence of hydramnios, side of herniation, cardiac deviation, stomach presence, gestational age at the time of finding the CDH and time of postpartum herniorrhaphy. The predictive values of these parameters for fetal outcome were analyzed. RESULTS: A total of 31 pregnancies were studied. There were 11 cases (35.5%) of termination, seven cases (22.6%) of perinatal death, four cases (12.9%) of late death and nine cases of survival (29%). The survivor group included four cases (44.4%) of complete recovery and five cases (55.6%) with persistent morbidity. There were 15 cases of simple CDH including eight cases of cardiac anomalies (ventricular-septal defect, atrial-septal defect, patent ductus arteriosus and ventricular dilatation). There were eight cases with severe anomalies (3 with trisomy 18, 2 with Cantrell's pentalogy, 1 with trisomy 13, 1 with cystic hygroma and one with Tetralogy Fallot). Among the seven parameters studied, gestational age at the time of finding the CDH and hydramnios were related to fetal survival. CONCLUSIONS: Sonography assists in predicting the postnatal outcome of CDH. Diagnosis of CDH at less than 25 weeks' gestation and the existence of hydramnios are associated with higher mortality. Postnatal therapy and prenatal surgical intervention are necessary to salvage fetuses in the presence of these two situations. The survival rate of infants with CDH was 45%. Of these, 55.6% had persistent morbidity. Prenatal counseling should reflect this.

Female↗

PubMed2000

Breech deformation complex in neonates.

OBJECTIVE: To demonstrate the association of minor anomalies and breech-presenting newborns with breech deformation complex. STUDY DESIGN: A total of 3,345 newborns with singleton, term delivery were examined based on a list of 67 items of major and minor anomalies. All infants were divided into two groups: group 1, vertex presentation (3,107 infants); group 2, breech presentation (224 infants). The prevalence of anomalies between both groups was compared. RESULTS: There were 1,495 (44.9%) infants with one or more minor anomalies, which included 1,313 infants (42.3%) in group 1 and 182 (81.3%) in group 2 (P < .005). The prevalence of anomalies in groups 1 and 2 were: frontal bossing (1.8% vs. 51%), prominent occiput (0.8% vs. 42%), upward slant (11.6% vs. 23.7%) and low-set ears (0.3% vs. 20.5%). Torticollis (0.3% vs. 1.78%) and congenital dislocation of the hip (CDH) (0.06% vs. 0.9%) (all P < .005) were related to breech presentation. CONCLUSION: Breech-presenting newborns had a higher risk of combining breech deformation complex, torticollis and CDH. The criteria for breech deformation complex included frontal bossing, prominent occiput, upward slant and low-set ears. The prevalence of breech deformation complex, torticollis and CDH was not related to the delivery method.

Abnormalities, Multiple↗

PubMed2000

Computed tomography and magnetic resonance imaging characteristics of giant pituitary adenomas.

PURPOSE: The purpose of this study was to characterize the computed tomography (CT) and magnetic resonance (MR) imaging features of giant pituitary adenoma (GPA) and to demonstrate the pathways of the spread of GPA. METHODS: CT and/or MR imaging scans of 356 patients evaluated preoperatively for single pituitary tumor were reviewed. Fourteen tumors (4%) fulfilled the radiologic criteria for GPA. There were 10 male and four female patients, ranging in age from 22 to 71 years (mean, 52 yr). We retrospectively reviewed the CT and MR imaging scans of these patients to characterize tumor appearances and identify the pathways of tumor extension. RESULTS: Thirteen tumors (93%) extended upward to the suprasellar cistern, and/or hypothalamus and third ventricle. Infrasellar extension through the sellar floor and sphenoid sinus to the skull base, or to the ethmoid sinus or the nasopharynx, was identified in seven patients (50%). Eight patients (57%) had lateral invasion to the cavernous sinus. Temporal and frontal extension was apparent in seven patients (50%) and six patients (43%), respectively. Five patients (36%) had posterior subtentorial extension to the posterior fossa. Histologically, only two GPAs showed invasive features. There was no correlation among histologic features, pituitary hormone concentrations, and evidence of tumor aggressiveness on CT and MRI scans. CONCLUSIONS: GPA has the potential for widespread, multi-directional extension. Our results indicate that any type of pituitary adenoma, regardless of its endocrinologic activity, may invade surrounding structures. Suprasellar invasion is the most common pathway of tumor spread, followed by infrasellar, lateral, anterior, and posterior routes.

Adenoma↗

PubMed1999

The midpoint potentials for the oxidized-semiquinone couple for Gly57 mutants of the Clostridium beijerinckii flavodoxin correlate with changes in the hydrogen-bonding interaction with the proton on N(5) of the reduced flavin mononucleotide cofactor as measured by NMR chemical shift temperature dependencies.

In the Clostridium beijerinckii flavodoxin, the reduction of the flavin mononucleotide (FMN) cofactor is accompanied by a local conformation change in which the Gly57-Asp58 peptide bond "flips" from primarily the unusual cis O-down conformation in the oxidized state to the trans O-up conformation such that a new hydrogen bond can be formed between the carbonyl group of Gly57 and the proton on N(5) of the neutral FMN semiquinone radical [Ludwig, M. L., Pattridge, K. A., Metzger, A. L., Dixon, M. M., Eren, M., Feng, Y., and Swenson, R. P. (1997) Biochemistry 36, 1259-1280]. This interaction is thought to contribute to the relative stabilization of the flavin semiquinone and may be at least partially responsible for the substantial separation of the midpoint potentials of the two one-electron reduction steps. Through a series of amino acid substitutions, the above cited study demonstrated the critical role of the often conserved glycine residue in this process. However, it has not been directly established experimentally as to whether these substitutions brought about the changes in the midpoint potentials by altering the strength of this hydrogen-bonding interaction as proposed. In this study, the relative strengths of the FMN N(5)H.O57 hydrogen bond in wild type and the G57A, G57N, and G57T mutants were evaluated by measuring the temperature dependency of the chemical shift for the proton on N(5) of the fully reduced cofactor by 1H-15N HSQC nuclear magnetic resonance spectroscopy. Based on the established correlation between the temperature coefficient of amide protons and the strength of hydrogen bonding in small peptides, the apparent strength of the N(5)H.O57 interaction was found to depend on the properties of the side chain at position 57. The glycine residue found in the wild-type flavodoxin appears to provide the strongest interaction while the beta-branched side chain in the G57T mutant provides the weakest. A good correlation was noted between the temperature coefficients of N(5)H and the one-electron reduction potential for the ox/sq couple as well as the binding free energy of the FMN semiquinone in this group of mutants. These results provide more direct quantitative evidence that support the previous hypothesis that this conformation change and the associated formation of the hydrogen bonding interaction with N(5)H of the reduced FMN represent an important means of stabilizing the neutral semiquinone and in modulating the oxidation-reduction potentials of the flavin cofactor in this and perhaps other flavodoxins.

Amino Acid Substitution↗

PubMed1999

Pituitary CRH receptor blockade reduces waking in the rat.

We have previously hypothesized that corticotropin-releasing hormone (CRH) is involved in the regulation of physiological waking. Central administration of CRH receptor antagonists reduces spontaneous waking in the rat. Some of the responses to central administration of CRH receptor antagonists may be mediated by mechanisms involving the hypothalamic-pituitary-adrenal axis, either by direct actions on the hypothalamus or by actions at the level of the pituitary due to leakage of the antagonists from the cerebrospinal fluid to blood. To further clarify the role of the hypothalamic-pituitary-adrenal axis as a mediator of responses to CRH receptor blockade, we administered intravenously into freely behaving rats in their home recording cages two specific CRH receptor antagonists, astressin or alpha-helical CRH, and determined subsequent changes in waking and sleep. Our results indicate that both antagonists reduce spontaneous waking, but with different time courses. Astressin, a potent antagonist of pituitary CRH receptors, reduces waking during postinjection hours 9-10, whereas high doses of alpha-helical CRH reduce waking only during the first postinjection hour. These results indicate that some effects of CRH on sleep-wake behavior may be meditated by pituitary CRH receptors.

Animals↗