Triple resonance-based assignment for Abl SH(32) and its complex with a consolidated ligand.
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Amino Acid Sequence↗
Biomedical subjects
S Cahill
Publications and source records attributed to S Cahill.
The solution structure and dynamics of the pleckstrin homology domain of G protein-coupled receptor kinase 2 (beta-adrenergic receptor kinase 1). A binding partner of Gbetagamma subunits.
The solution structure of an extended pleckstrin homology (PH) domain from the beta-adrenergic receptor kinase is obtained by high resolution NMR. The structure establishes that the beta-adrenergic receptor kinase extended PH domain has the same fold and topology as other PH domains, and there are several unique features, most notably an extended C-terminal alpha-helix that behaves as a molten helix, and a surface charge polarity that is extensively modified by positive residues in the extended alpha-helix and the C terminus. These observations complement biochemical evidence that the C-terminal portion of this PH domain participates in protein-protein interactions with Gbetagamma subunits. This suggests that the C-terminal segment of the PH domain may function to mediate protein-protein interactions with the targets of PH domains.
Amino Acid Sequence↗
The main-chain dynamics of the dynamin pleckstrin homology (PH) domain in solution: analysis of 15N relaxation with monomer/dimer equilibration.
The backbone dynamics of the pleckstrin homology (PH) domain from dynamin were studied by 15N NMR relaxation (R1 and R2) and steady state heteronuclear 15N [1H] nuclear Overhauser effect measurements at 500 and 600 MHz, at protein concentrations of 1.7 mM and 300 microM, and by molecular dynamics (MD) simulations. The analysis was performed using the model-free approach. The method was extended in order to account for observed partial (equilibrium) dimerization of the protein at NMR concentrations. A model is developed that takes into account both rapid monomer-dimer exchange and anisotropy of the over-all rotation of the dimer. The data show complex dynamics of the dynamin PH domain. Internal motions in elements of the secondary structure are restricted, as inferred from the high value of the order parameter (S2 approximately 0.9) and from the local correlation time < 100 ps. Of the four extended loop regions that are disordered in the NMR-derived solution structure of the protein, loops beta 1/beta 2 and beta 5/beta 6 are involved in a large-amplitude (S2 down to 0.2 to 0.3) subnanosecond to nanosecond time-scale motion. Reorientation of the loops beta 3/beta 4 and beta 6/beta 7, in contrast, is restricted, characterized by the values of order parameter S2 approximately 0.9 more typical of the protein core. These loops, however, are involved in much slower processes of motion resulting in a conformational exchange on a microsecond to submillisecond time scale. The motions of the terminal regions (residues 1 to 10, 122 to 125) are practically unrestricted (S2 down to 0.05, characteristic times in nanosecond time scale), suggesting that these parts of the sequence do not participate in the protein fold. The analysis shows a larger sensitivity of the 15N relaxation data to protein microdynamic parameters (S2, tau loc) when protein molecular mass (tau c) increases. The use of negative values of the steady state 15N[1H] NOEs as an indicator of the residues not belonging to the folded structure is suggested. The amplitudes of local motion observed in the MD simulation are in a good-agreement with the NMR data for the amide NH groups located in the protein core.
Blood Proteins↗
Condom use in a group of aboriginal women.
This exploratory study investigated the attitudes of Aboriginal women in Darwin to the use of condoms to prevent HIV and other sexually transmitted diseases. An Aboriginal research assistant interviewed twelve women regarding their usage of and attitudes to the male condom and their attitude to the possibility of using a female condom. These women, like their ethnic minority counterparts in Africa and North America, had a fair level of knowledge about HIV, a low level of perceived risk of HIV and other sexually transmitted diseases, infrequent usage of male condoms with their partner, and negative attitudes to the male condom. Their attitude to the female condom was more positive. Educational programs and further research into the attitudes of women in remote communities are recommended.
Adult↗
Ductal carcinoma in situ. Part 2: Treatment.
Several dilemmas exist when treating a patient with ductal carcinoma in situ (DCIS): the high rate of inter-observer variation for pathologists who must diagnose these tumours; the potential for over- and under-treatment; and the uncertainty about the best way to inform a patient who must often make a decision between breast conservation and mastectomy. Mastectomy is nearly 100% curative, is expedient, but may represent over-treatment for many women, particularly those with asymptomatic mammographically detected lesions. Axillary dissection is not recommended as a routine except for patients with lesions over 5 cm in whom the risk of micro-invasion and lymph node involvement increases. Conservative surgery (CS) alone is associated with a local recurrence rate of approximately 20%, and half of these recurrences (10% overall) are invasive, with a potential long-term cure rate of at least 90%. The addition of radiation to CS reduces the risk of local recurrence to approximately 10%, half of these recurrences (5%) are invasive for a potential long-term cure rate of 95%. Several randomized trials comparing CS with or without radiation therapy (RT) are in progress. The factors that increase the rate of local recurrence after CS alone for DCIS include close or involved margins, and the presence of necrosis or high-grade tumours. Patients with these features should have radiation therapy if breast conservation is preferred. Patients with low-grade tumours (without necrosis) up to 15 mm, with clear margins of at least 10 mm, who agree to be closely observed may be good candidates for CS alone. A critical review of the literature is presented.
Breast Neoplasms↗
The behavior of the active site salt bridge of bovine neurophysins as monitored by 15N NMR spectroscopy and chemical substitution. Relationship to biochemical properties.
The active site of liganded neurophysin contains a salt bridge that involves the side chains of Arg-8 and Glu-47 of the protein and the alpha-amino group of bound hormone or related peptide. The extent to which the Arg-8-Glu-47 salt bridge persists in the absence of peptide, or to which the environment of Arg-8 in the unliganded state differs in monomers and dimers, is relevant to an understanding of allosteric mechanism in this system. In the present study, the behavior of the salt bridge was investigated by 15N NMR and chemical replacement of Arg-8. Bovine neurophysin-I was converted to its des 1-8 derivative, and Arg-8 was replaced by 15N-substituted Arg or by other residues using chemical semisynthesis. The relative abilities of different amino acids to restore peptide affinity to the des 1-8 protein were in good accord with the view of the salt bridge in the liganded state obtained from crystals of bovine neurophysin-II complexes. In the unliganded state, comparison of the 15N and proton NMR signals from Arg-8 with those in smaller arginine systems suggested the absence of significant interactions between the guanidinium of Arg-8 and Glu-47 or between the amino terminal region of Arg-8 and other elements of the protein. No evidence of a difference in Arg-8 environment between unliganded monomers and dimers was found. Marked spectral changes accompanying the binding of oxytocin indicated changes in the environment of both the side chain and amino terminal region of Arg-8. The NMR results were in good agreement with a recently emerging comparison of bovine neurophysin-II derivatives in the liganded and unliganded states, with the notable exception of the extent of salt bridge formation in the unliganded state. The results are shown to be consistent with, and to help explain, significant differences between the two bovine neurophysins in the susceptibility to tryptic cleavage at Arg-8 in the unliganded state and in the pH dependence of peptide binding and additionally constrain potential allosteric mechanisms underlying neurophysin ligand-facilitated dimerization.
Allosteric Regulation↗
Solution structure of pleckstrin homology domain of dynamin by heteronuclear NMR spectroscopy.
The pleckstrin homology (PH) domain is a recognition motif thought to be involved in signal-transduction pathways controlled by a variety of cytoplasmic proteins. Assignments of nearly all 1H, 13C, and 15N resonances of the PH domain from dynamin have been obtained from homonuclear and heteronuclear NMR experiments. The secondary structure has been elucidated from the pattern of nuclear Overhauser enhancements, from 13C chemical shift deviations, and from observation of slowly exchanging amide hydrogens. The secondary structure contains one alpha-helix and eight beta-strands, seven of which are arranged in two contiguous, antiparallel beta-sheets. The structure is monomeric, in contrast to the well-defined intimate dimerization of the crystal structure of this molecule. Residues possibly involved in ligand binding are in apparently flexible loops. Steady-state 15N(1H) nuclear Overhauser effect measurements indicate unequivocally the boundaries of this PH domain, and the structured portion of the domain appears to be more extended to the C terminus than previously suggested for other PH domains.
Amino Acid Sequence↗
Vasoactive intestinal peptide and nerve growth factor effects on nerve regeneration.
Sixty rat sciatic nerves were used to study the effects of vasoactive intestinal peptide (VIP) and nerve growth factor (NGF) on nerve regeneration. They were divided into three groups. Groups 1 and 2 were treated with VIP and NGF, respectively, after dividing the nerves without repair and placing them in silastic chambers. Group 3 served a control. The rate and quality of nerve regeneration were compared among the groups using caliper measurements and histologic evaluation. Both VIP and NGF groups showed an enhanced rate of regeneration at three weeks as compared to controls (p < 0.05). The quality of nerve regeneration histologically and by axonal counting was not significantly different among the three groups, except for the presence of less vascularity in the VIP as compared with the NGF group. In addition to NGF, VIP appears to increase the rate of nerve regeneration as compared to controls.
Animals↗
Properties of a recombinant human hemoglobin with aspartic acid 99(beta), an important intersubunit contact site, substituted by lysine.
Site-directed mutagenesis of an important subunit contact site, Asp-99(beta), by a Lys residue (D99K(beta)) was proven by sequencing the entire beta-globin gene and the mutant tryptic peptide. Oxygen equilibrium curves of the mutant hemoglobin (Hb) (2-15 mM in heme) indicated that it had an increased oxygen affinity and a lowered but significant amount of cooperativity compared to native HbA. However, in contrast to normal HbA, oxygen binding of the recombinant mutant Hb was only marginally affected by the allosteric regulators 2,3-diphosphoglycerate or inositol hexaphosphate and was not at all responsive to chloride. The efficiency of oxygen binding by HbA in the presence of allosteric regulators was limited by the mutant Hb. At concentrations of 0.2 mM or lower in heme, the mutant D99K(beta) Hb was predominantly a dimer as demonstrated by gel filtration, haptoglobin binding, fluorescence quenching, and light scattering. The purified dimeric recombinant Hb mutant exists in 2 forms that are separable on isoelectric focusing by about 0.1 pH unit, in contrast to tetrameric hemoglobin, which shows 1 band. These mutant forms, which were present in a ratio of 60:40, had the same masses for their heme and globin moieties as determined by mass spectrometry. The elution positions of the alpha- and beta-globin subunits on HPLC were identical. Circular dichroism studies showed that one form of the mutant Hb had a negative ellipticity at 410 nm and the other had positive ellipticity at this wavelength. The findings suggest that the 2 D99K(beta) recombinant mutant forms have differences in their heme-protein environments.
Amino Acid Sequence↗
Vesicoureteral reflux: an accurate predictor of acute pyelonephritis in childhood urinary tract infection?
PURPOSE: To determine the association between vesicoureteral reflux (VUR) and the presence of acute pyelonephritis in children with urinary tract infections. MATERIALS AND METHODS: The authors studied 150 consecutive patients less than 5 years of age with their first proved urinary tract infection. All patients underwent renal cortical scintigraphy (with technetium-99m dimercaptosuccinic acid or Tc-99m gluconate) and voiding cystourethrography (VCUG) to identify the presence of cortical defects and VUR, respectively. RESULTS: Of 300 kidneys, 88 (29.3%) had a cortical defect at scintigraphy. Fifty-four of the 88 patients (61%) did not have VUR demonstrated at VCUG. Conversely, 72 of the 300 kidneys (24%) had VUR; of these, 38 (53%) had no cortical defect. The sensitivity of VCUG in helping predict a defect was 38.6%, and the specificity was 82.1%. CONCLUSION: VUR (as shown by VCUG) and renal cortical scintigraphic defects frequently occur independently of each other. Renal cortical scintigraphy may be a more accurate predictor of patients at risk for scarring.
Acute Disease↗
Risk factors in the development of early renal cortical defects in children with urinary tract infection.
OBJECTIVE: Defects seen on early cortical scintigrams of the renal cortex in children with urinary tract infection may represent acute inflammatory change or established scar. The purpose of this study was to determine the relationship between these defects and age, sex, the presence and grade of vesicoureteral reflux, and infective organism in a cohort of children examined after their first proved urinary tract infection. SUBJECTS AND METHODS: We prospectively examined 193 consecutive patients less than 5 years old who were seen at the ambulatory pediatric department during a 3-year period and had a first proved urinary tract infection. Children with obstructed or solitary kidneys were excluded. All patients were imaged with scintigraphy of the renal cortex and radiographic voiding cystourethrography within 15 days of diagnosis. The association of age, sex, the presence and grade of vesicoureteral reflux, and infective organism with a defect (acute pyelonephritis or a renal scar) seen on a cortical renal scan was studied. RESULTS: The prevalence of cortical defects was greater in the kidneys of patients less than 2 years old (96/290, 33%) than in older children (16/96, 17%) and greater in those with vesicoureteral reflux (41/92, 45%) than in those without it (71/294, 24%). Vesicoureteral reflux was absent in 63% (71/112) of kidneys with a cortical defect. No association with sex or infective organism was established. As well as having a greater prevalence of cortical defects, 145 (75%) of the 193 urinary tract infections included in the study were in children less than 2 years old. The kidneys of these younger patients also had a greater severity and prevalence of vesicoureteral reflux (74/290, 26%) than did those of older children (18/96, 19%). CONCLUSION: Early cortical defects are associated with an age less than 2 years and vesicoureteral reflux. However, the association of early defects with the presence and grade of vesicoureteral reflux is confounded by the declining prevalence and severity of reflux with age. A significant proportion of cortical defects occur in the absence of vesicoureteral reflux, and the contribution of reflux to scar formation might be less than previously considered.
Age Distribution↗
Role of bipedal lymphangiogram in radiation treatment planning for cervix cancer.
PURPOSE: The role of bipedal lymphangiogram in tailoring radiation portals in radical radiation therapy for Stages II or III cervix cancer is investigated. METHODS AND MATERIALS: The records and simulation films of 87 patients with Stage II or III carcinoma of the cervix treated with radical radiation therapy alone have been retrospectively reviewed. RESULTS: Sixty-two percent of patients who had a bipedal lymphangiogram, subsequently had their radiation fields altered from a "standard portal." The most frequently altered fields were the lateral margin of the postero anterior field and the anterior margin of the lateral fields. In order to cover the lymphatic channels in the pelvis in 90% of cases as outlined by the bipedal lymphangiogram, the lateral margins of the postero anterior fields would need to be 2.5 cm lateral to the pelvic brim and the anterior border of the lateral field, 0.5 cm anterior to the pubic symphysis. CONCLUSION: "Standard" fields adequately cover the median distribution of lymphatics, but may result in a geographic miss in some patients. Bipedal lymphangiograms allow a more accurate tailoring of pelvic lymphatic fields.
Australia↗
Effects of rapid pulsed CO2 laser beam on cortical bone in vivo.
Twenty rabbit femurs were used to study the effect of CO2 laser on cortical bone. Sixteen femurs were treated with 20 watts, 3 mm defocused beam, 2 KHZ spike pulse mode CO2 laser for 10 seconds through a circular window in the metaphysis. In four control femurs, the inner cortex was exposed without laser treatment. The animals were killed at 4 and 6 weeks and the specimens studied histologically. All laser-treated specimens showed thermal changes. Three histological zones were observed. A superficial zone of inner cortex close to the beam consisted mainly of carbonization or carbon ash during resorption. An intermediate zone consisted of bone necrosis and healing with associated areas of new bone formation. The deep zone of outer cortex had normal bone with no cellular damage. No such changes were observed in the control specimens. The CO2 laser can be used to generate a controlled zone of tissue ablation, which may make it a potentially useful tool for tumor margin cauterization.
Animals↗
Comparison of DMSA scintigraphy with intravenous urography for the detection of renal scarring and its correlation with vesicoureteric reflux.
A series of 208 patients was prospectively assessed for reflux nephropathy by intravenous urography (IVU) and 99mTc-dimercaptosuccinate (DMSA) scintigraphy. All patients were studied at least 3 months after their most recent urinary tract infection and micturating cystourethrography (MCU) was performed prior to the scintigraphic studies. DMSA scintigraphy detected significantly more cortical abnormalities than did IVU. There was also a correlation between cortical abnormalities in the DMSA studies and the degree of reflux on MCU. The validity of DMSA as a cortical imaging agent is evaluated and the histological evidence for its efficacy derived from the animal model is reviewed, lending weight to its establishment as the "gold standard" for renal cortical scarring.
Child↗
A comparison of human and animal mouth flora.
Human bite injuries are often believed to be more serious than animal bite injuries. One reason for this distinction might be a species difference in the microbial mouth flora. In order to compare this possibility, we cultured the mouths of humans, dogs, and cats. Human oral flora contained the smallest number of bacteria followed by dog and cat oral flora, respectively. Veillonella parvula was cultured from all humans and was also found in dog and cat mouths, but less frequently. V parvula is not usually considered a virulent organism but occasionally may become a pathogen and should not be overlooked especially following human bite injury.
Animals↗
DMSA studies in infants under one year of age.
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Age Factors↗
The relation of social class to risk factors, rehabilitation, compliance and mortality in survivors of acute coronary heart disease.
We studied 299 consecutive male 28-day survivors of unstable angina or myocardial infarction aged under 60 years to examine the relationship between social class and initial risk factors, change in risk-factors at one year follow-up, return to work, and 3-year mortality. There was a significant correlation between smoking on admission and social class, with 80% of lower and 31% of upper classes being current smokers. Daily cigarette consumption among smokers was significantly higher in lower-class patients. Lower-class patients also had a significantly higher weekly alcohol intake. Although the proportion of hypertensives did not vary with social class, mean in-hospital blood pressure was higher in lower-class patients. Social class bore no relationship to amount of leisure exercise, serum cholesterol or degree of overweight. There was a 90% 1-year return to work overall, and while there was no relationship between social class and eventual re-employment, lower-class patients took significantly longer to return to work. There were highly significant associations between social class and successful smoking cessation, increase in leisure exercise and weight reduction over the first year after discharge. There was no significant association between social class and 3-year mortality.
Angina, Unstable↗
An immunochemical approach to the identification of the MBTA binding site of the nicotinic acetylcholine receptor of Torpedo californica.
Monospecific anti-[4-(N-maleimidobenzyl) trimethylammonium] (MBTA) antibodies were prepared from sera of rabbits immunized with an albumin-MBTA conjugate and used to synthesize an MBTA-specific immunosorbent. Torpedo californica acetylcholine receptor was affinity labeled with [3H]-MBTA and proteolyzed extensively with pronase, and the peptide fraction of the digest chromatographed on the anti-MBTA resin. The amino acid composition of the purified MBTA-peptide fraction was compared with the sequences flanking the seven cysteinyl residues of the alpha-subunit. The best fit was observed with the segment containing cysteine 142.
Affinity Labels↗