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Biomedical subjects

D O'Connell

Publications and source records attributed to D O'Connell.

At least 19 recordsLinked to original sources
PubMed2003

Platelet-rich-plasmapheresis for minimising peri-operative allogeneic blood transfusion.

BACKGROUND: Concerns regarding the safety of transfused blood have generated considerable enthusiasm for the use of technologies intended to reduce the use of allogeneic blood (blood from an unrelated donor). Platelet-rich plasmapheresis (PRP) offers an alternative approach to blood conservation. OBJECTIVES: To examine the evidence for the efficacy of PRP in reducing peri-operative allogeneic red blood cell (RBC) transfusion, and the evidence for any effect on clinical outcomes such as mortality and re-operation rates. SEARCH STRATEGY: Studies were identified by: computer searches of MEDLINE, EMBASE, Current Contents, and the Cochrane Library (to June 2001). These searches were supplemented by checking the reference lists of published articles, reports, and reviews. SELECTION CRITERIA: Controlled parallel group trials in which adult patients, scheduled for non-urgent surgery, were randomised to PRP, or to a control group who did not receive the intervention. DATA COLLECTION AND ANALYSIS: Main outcomes measured were: the number of patients receiving an allogeneic RBC transfusion, and the amount of RBC transfused. Trial quality was assessed using criteria proposed by Schulz et al. (Schulz 1995) and Jadad et al. (Jadad 1996). MAIN RESULTS: Nineteen trials of PRP were identified that reported data for the number of patients exposed to allogeneic RBC transfusion. These trials evaluated a total of 1452 patients. The pooled relative risk (RR) of exposure to allogeneic blood transfusion in those patients randomised to PRP was 0.71 (95%CI: 0.56, 0.90), equating to a relative risk reduction (RRR) of 29%; the average absolute risk reduction (ARR) was 19% (RD = -0.19: 95%CI: -0.29, -0.09). On average, PRP did not significantly reduce the total volume of RBC transfused (weighted mean difference [WMD] = -0.69: 95%CI: -1.93, 0.56 units). Substantial statistical heterogeneity was observed (p < 0.001). Trials provided inadequate data regarding the impact of PRP on morbidity, mortality, and hospital length of stay. The majority of trials were small and of poor methodological quality. REVIEWER'S CONCLUSIONS: Although the results suggest that PRP is effective in reducing allogeneic RBC transfusion in adult patients undergoing elective surgery, there was considerable heterogeneity in treatment effects and the trials were of poor methodological quality. As the majority of trials were unblinded, transfusion practices may have been influenced by knowledge of the patient's allocation status, potentially exaggerating the true magnitude of the beneficial effect of PRP. The available studies provided inadequate data for firm conclusions to be drawn regarding the impact of PRP on clinically important endpoints.

Adult↗
PubMed2002

Pre-operative autologous donation for minimising perioperative allogeneic blood transfusion.

BACKGROUND: Public concerns regarding the safety of transfused blood have prompted re-consideration of the indications for the transfusion of allogeneic red cells (blood from an unrelated donor), and a range of techniques designed to minimise transfusion requirements. OBJECTIVES: To examine the evidence for the efficacy of pre-operative autologous blood donation (PAD) in reducing the need for peri-operative allogeneic red blood cell (RBC) transfusion. SEARCH STRATEGY: Articles were identified by: computer searches of OVID MEDLINE, EMBASE, and Current Contents (to March 2001) and web sites of international health technology assessment agencies (to January 2001). References in the identified trials were checked and authors contacted to identify additional studies. SELECTION CRITERIA: Randomised controlled trials with a concurrent control group in which adult patients, scheduled for non-urgent surgery, were randomised to PAD, or to a control group who did not receive the intervention. DATA COLLECTION AND ANALYSIS: Trial quality was assessed using criteria proposed by Schulz et al (1995) and Jadad et al (1996). The principle outcomes were: the number of patients exposed to allogeneic red blood cells, and the amount of blood transfused. Other clinical outcomes are detailed in the review. MAIN RESULTS: Overall PAD reduced the risk of receiving an allogeneic blood transfusion by a relative 63% (RR=0.37: 95%CI:0.26,0.54). The absolute reduction in risk of allogeneic transfusion was 43.8% (RD=-0.438: 95%CI: -0.607,-0.268). In contrast the results show that the risk of receiving any blood transfusion (allogeneic and/or autologous) is actually increased by pre-operative autologous blood donation (RR=1.29: 95%CI: 1.12,1.48). Trials were unblinded and allocation concealment was not described in 87.5% of the trials. REVIEWER'S CONCLUSIONS: Although the trials of PAD showed a reduction in the need for allogeneic blood the methodological quality of the trials was poor and the overall transfusion rates (allogeneic and/or autologous) in these trials were high, and were increased by recruitment into the PAD arms of the trials. This raises questions about the true benefit of PAD. In the absence of large, high quality trials using clinical endpoints, it is not possible to say whether the benefits of PAD outweigh the harms.

Blood Transfusion, Autologous↗
PubMed2001

Desmopressin for minimising perioperative allogeneic blood transfusion.

BACKGROUND: Public concerns regarding the safety of transfused blood have prompted re-consideration of the use of allogeneic (from an unrelated donor) red blood cell (RBC) transfusion, and a range of techniques designed to minimise transfusion requirements. OBJECTIVES: To examine the evidence for the efficacy of desmopressin (1-deamino-8-D-arginine-vasopressin), in reducing perioperative blood loss and the need for red cell transfusion in patients who do not have congenital bleeding disorders. SEARCH STRATEGY: Articles were identified by: computer searches of OVID MEDLINE, EMBASE, and Current Contents (to August 2000) and web sites of international health technology assessment agencies (to May 1998). References in the identified trials and review articles were checked and authors contacted to identify additional studies. SELECTION CRITERIA: Randomised controlled parallel group trials in which adult patients, scheduled for non-urgent surgery, were randomised to DDAVP, or to a control group, who did not receive the intervention. DATA COLLECTION AND ANALYSIS: Trial quality was assessed using criteria proposed by Schulz et al. (1995) and Jadad et al. (1996). The principal outcomes were: the number of patients exposed to red cells, and the amount of blood transfused. Other clinical outcomes are detailed in the review. MAIN RESULTS: Fourteen trials of DDAVP (N=1034) reported data on the proportion of patients exposed to allogeneic RBC transfusion. In subjects treated with DDAVP the relative risk of exposure to peri-operative allogeneic blood transfusion was 0.98 (95%CI: 0.88 to 1.10) compared with control. In DDAVP-treated patients the relative risk of requiring re-operation due to bleeding was 0.56 (95%CI: 0.18 to 1.73). There was no statistically significant effect overall for mortality and non-fatal myocardial infarction in DDAVP-treated patients compared with control (RR=1.53: 95%CI: 0.58 to 4.05) and (RR=1.52: 95%CI: 0.67 to 3.49) respectively. REVIEWER'S CONCLUSIONS: There is no convincing evidence that desmopressin minimises perioperative allogeneic RBC transfusion in patients who do not have congenital bleeding disorders. These data suggest that there is no benefit of using DDAVP as a means of minimising perioperative allogeneic RBC transfusion. This meta-analysis had 90% power to detect a relative risk reduction of at least 17% for receiving a red cell transfusion at alpha = 0.05 (two-sided).

Adult↗
PubMed2001

Anti-fibrinolytic use for minimising perioperative allogeneic blood transfusion.

BACKGROUND: Concerns regarding the safety of transfused blood have prompted re-consideration of the use of allogeneic (blood from an unrelated donor) blood transfusion. OBJECTIVES: To assess the effects of the anti-fibrinolytic drugs aprotinin, tranexamic acid, and epsilon aminocaproic acid, on peri-operative red blood cell (RBC) transfusion. SEARCH STRATEGY: We searched MEDLINE (to May 1998), EMBASE (to December 1997), web sites of international health technology assessment agencies (to May 1998). References in identified trials and review articles were checked and authors contacted to identify any additional studies. SELECTION CRITERIA: Randomised controlled trials of anti-fibrinolytic drugs in adults scheduled for non-urgent surgery. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality and extracted data. MAIN RESULTS: We found 61 trials of aprotinin (7027 participants). Aprotinin reduced the rate of RBC transfusion by a relative 30% (RR=0.70: 95%CI: 0.64 to 0.76). The average absolute risk reduction (ARR) was 20.4% (95%CI: 15.6% to 25.3%). On average, aprotinin use saved 1.1 units of RBC (95%CI: 0.69 to 1.47) in those requiring transfusion. Aprotinin also significantly reduced the need for re-operation due to bleeding (RR=0.40: 95%CI: 0.25 to 0.66). We found 18 trials of tranexamic acid (TXA) (1,342 participants). TXA reduced the rate of RBC transfusion by a relative 34% (RR=0.66: 95%CI: 0.54 to 0.81). This represented an ARR of 17.2% (95%CI: 8.7% to 25.7%). TXA use resulted in a saving of 1.03 units of RBC (95%CI: 0.67 to 1.39) in those requiring transfusion. We found four trials of epsilon aminocaproic acid (EACA) (208 participants). EACA use resulted in a statistically non-significant reduction in RBC transfusion (RR=0.48: 95%CI: 0.19 to 1.19). Comparisons between agents Eight trials made 'head-to-head' comparisons between TXA and aprotinin. There was no significant difference between the two drugs in the rate of RBC transfusion: RR=1.21 (95%CI: 0.83 to 1.76) for TXA compared to aprotinin. Adverse Effects Aprotinin did not seem to be associated with an excess risk of adverse effects, including thrombo-embolic events (thrombosis RR=0.64: 95%CI: 0.31 to 1.31) and renal failure (RR=1.19: 95%CI: 0.79 to 1.79). Fewer data were available for TXA and EACA. REVIEWER'S CONCLUSIONS: From this review it appears that aprotinin reduces the need for red cell transfusion, and the need for re-operation due to bleeding, without serious adverse effects. However, there was significant heterogeneity in trial outcomes, and some evidence of publication bias. Similar trends were seen with TXA and EACA, although the data were rather sparse. The poor evaluation of these latter drugs is unfortunate as results suggest they may be equally as effective as aprotinin, but are significantly cheaper. The evidence reviewed here supports the use of aprotinin in cardiac surgery. Further small trials of this drug are not warranted. Future trials should be large enough to compare the efficacy and cost-effectiveness of aprotinin with that of TXA and EACA.

Aminocaproic Acid↗
PubMed2001

The effects of fenoldopam on coronary conduit blood flow after coronary artery bypass graft surgery.

OBJECTIVE: To quantify the effects of fenoldopam, 0.1 microg/kg/min, on left internal mammary artery (LIMA) and saphenous vein blood flow after coronary anastomosis. DESIGN: Prospective, randomized, double-blind, placebo-controlled trial. SETTING: University teaching hospital, single institution. PARTICIPANTS: Thirty-one American Society of Anesthesiologists III patients undergoing elective coronary revascularization. INTERVENTIONS: A perivascular ultrasonic flow probe (Linton Instrumentation, Norfolk, UK) was placed around the LIMA and saphenous vein graft after coronary anastomosis. MEASUREMENTS AND MAIN RESULTS: Immediately before and at 5-minute intervals for 15 minutes after starting the infusion, blood flow was measured in the LIMA and one saphenous vein graft using a transit time ultrasonic flow probe. Heart rate, blood pressure, and central venous pressure were documented at these time points. Administration of fenoldopam, 0.1 microg/kg/min, did not alter heart rate or blood pressure. A small, nonsignificant increase in LIMA blood flow occurred during the 15-minute study period (30 +/- 12 to 35 +/- 10 mL/min) in patients who received fenoldopam. No significant changes occurred in the placebo group. CONCLUSIONS: The findings indicate that fenoldopam, 0.1 microg/kg/min, did not influence coronary conduit blood flow to a clinically significant extent. The small increase in LIMA blood flow may be of greater importance in high-risk patients or in the prevention of coronary arterial spasm.

Aged↗
PubMed2000

A role for the integrin alphavbeta8 in the negative regulation of epithelial cell growth.

The control of cell growth is regulated through coordinated responses to growth factors and cell-extracellular matrix (ECM) interactions. Integrins, the major family of cell-ECM receptors, are vital to these coordinated responses. Although much is known of the role of integrins in growth promotion, specific examples of integrin-mediated cell growth inhibition are few. On the basis of our findings that the integrin beta8 subunit is expressed in airway epithelial cells and is absent in lung cancers, we investigated the role and mechanism of the integrin alphavbeta8 in mediating growth inhibition. When introduced into either a lung or colon carcinoma cell line, beta8 inhibited cell growth without inducing apoptosis. Ligation of alphavbeta8 also induced cell rounding, inhibited focal contact formation, and initiated an inhibitory signaling pathway as demonstrated by increased expression of the cyclin-dependent kinase inhibitor p21Cip1. The cytoplasmic domain of beta8 was capable of both growth inhibition and causing cell shape changes as shown by the use of a chimeric integrin construct consisting of the beta8-cytoplasmic domain coupled to the beta6-extracellular domain. Finally, when tested in vivo, beta8 potently inhibited tumor growth in nude mice. Together, these results implicate alphavbeta8 as a novel growth-regulatory molecule of epithelial cells.

Actins↗
PubMed2000

Tricyclic drugs for depression in children and adolescents.

BACKGROUND: There is a need to identify effective and safe treatments for depression in children and adolescents. While tricyclic drugs have proven effectiveness in the treatment of depression in adults, individual studies involving children and adolescent have been equivocal. OBJECTIVES: To examine whether orally administered tricyclic antidepressants are superior to placebo in the treatment of child and adolescent depression SEARCH STRATEGY: We searched the literature using CD ROM Silver Platter and On-Line MEDLINE (1966-1997) and Excerpta Medica (June 1974-1997) data bases. Terms used for the search were: the exploded terms child and depression; the Medical Subject Headings of antidepressant drugs, tricyclic and affective disorders; individual tricyclic drugs by name; names of well-known researchers in the field; and school phobia. We searched the trials database of the Cochrane Collaboration Depression, Anxiety and Neurosis Group. Abstracts in English (of English and non-English papers) were reviewed. Bibliographies of previously published reviews and papers describing original research were cross-checked. Current Contents was screened for recent publications. We contacted authors of abstracts describing "work in progress" identified in conference proceedings of the American Academy of Child and Adolescent Psychiatry to determine whether they held data which could be included in the meta-analysis. We have hand searched the Journal of the American Academy of Child and Adolescent Psychiatry to identify randomized controlled trials. SELECTION CRITERIA: Randomized controlled trials comparing the efficacy of orally administered tricyclic medication with placebo in depressed subjects aged 6-18 years. DATA COLLECTION AND ANALYSIS: Most studies reported multiple outcome measures including depression scales and clinical global impression scales. For each study the best available depression measure was taken as the index measure of depression outcome. Predetermined criteria were established to assist in the ranking of measures. Where authors reported categorical outcomes we calculated individual and pooled odds ratios for the odds of improvement in treated compared with control subjects. For continuous outcomes pooled effect sizes were calculated as the number of standard deviations by which the change in depression scores for the treatment group exceeded those for the control groups. MAIN RESULTS: Twelve studies fulfilled the criteria for inclusion in the review, eight from which dichotomous outcome data could be extracted, and eleven from which continuous outcome data could be extracted. Pooled odds ratios calculated from the dichotomous data indicated no advantage of treatment over placebo for children or adolescents (odds ratio = 0.83, 95% confidence interval 0.48 to 1.42). Effect size calculations from the continuous data suggested a statistically significant but small benefit of treatment over placebo in reducing symptoms for the aggregate sample (effect size = -0.38, 95% confidence interval -0.74 to -0.02) with subgroup analyses suggesting a larger benefit among adolescents (effect size = -0.59, 95% confidence interval -1.12 to -0.06), and no benefit among children (effect size = 0.15, 95% confidence interval -0.34 to 0.64). Treatment with a tricyclic antidepressant caused more vertigo (odds ratio = 8.47, 95% confidence interval 1.40 to 51.0), orthostatic hypotension (odds ratio = 4.77, 95% confidence interval 1.11 to 20.49) and dry mouth (odds ratio = 5.19, 95% confidence interval 1.15 to 23.51) than did placebo, but there was no statistically significant difference in other possible adverse effects. REVIEWER'S CONCLUSIONS: Data suggest tricyclic antidepressants are of unlikely benefit in the treatment of depression in pre pubertal children. There is marginal evidence to support the use of tricyclic antidepressants in the treatment of depression in adolescents, although the magnitude of effect is likely to be moderate at best.

Administration, Oral↗
PubMed2000

Giant distal humeral geode.

We describe the imaging features of a giant geode of the distal humerus in a patient with rheumatoid arthritis, which presented initially as a pathological fracture. The value of magnetic resonance imaging in establishing this diagnosis is emphasized.

Arthritis, Rheumatoid↗
PubMed2000

Rapid analysis of illicit drugs by mass spectrometry: results from seizures in Ireland.

A gas chromatographic procedure with mass spectrometric detection (GC-MS) was established to determine the principal amphetamines, methylenedioxymethamphetamine (MDMA), methylenedioxyethamphetamine (MDEA) and methylenedioxyamphetamine (MDA), cocaine and pharmacologically active impurities in 'ecstasy' tablets. The procedure was developed and optimised by combining the individual methods for the various drugs of abuse available in the literature into a single GC-MS run. New variants of the main drugs which may appear on the drug scene were also detected, including N-methyl-1-phenylethylamine, a member of a new series of illicit drugs. The method is rapid and has good sensitivity and reproducibility.

Amphetamines↗
PubMed2000

Anti-L-selectin aptamers: binding characteristics, pharmacokinetic parameters, and activity against an intravascular target in vivo.

Therapeutic and diagnostic applications have been envisioned for aptamers, a class of oligonucleotide ligands that bind their target molecules with high affinity and specificity (Gold, J. Biol. Chem. 270, 13581-13584, 1995). To identify parameters that are important for the in vivo activity of aptamers acting on intravascular targets, we have studied binding characteristics in vitro, pharmacokinetic parameters in Sprague-Dawley rats, and inhibitory activity in a SCID mouse/human lymphocyte model of lymphocyte trafficking for both 2'F pyrimidine 2'OH purine RNA and ssDNA anti-human L-selectin aptamers. The data indicate that aptamers with low nanomolar affinity are suitable candidates for use as in vivo reagents and that nonspecific binding to vascular cells is not an issue for efficacy. As is often observed for other reagents, plasma clearance is biphasic. Both the distribution phase and the clearance rate strongly affect in vivo activity. Pharmacokinetic parameters and in vivo activity are significantly improved by conjugating aptamers to a carrier molecule, such as polyethylene glycol (PEG). Most active in vivo is 1d40, a 2'F pyrimidine 2'OH purine aptamer conjugated to 40 kDa PEG. At a dose of 5.4 nmol/kg body weight, its duration of effect (time to 50% inhibition) is 11.2 hours, and at 1 mg or 90 nmol/kg, its plasma clearance rate (CL) is 0.4 ml/min/kg. Its ED50 is estimated to be 80 pmol/kg in preinjection dose-response experiments, compared with 4 pmol/kg for the dimeric anti-L-selectin antibody DREG56. Further improvement of in vivo activity is expected from nucleotide modifications that increase resistance to nuclease digestion for aptamers where mass is not rate limiting for clearance. Because the relationship of clearance to conjugate molecular weight (MW) is not the same for all aptamers, it is advisable to determine the relationship at the outset of in vivo studies. In summary, the data suggest that properly formulated aptamers have the capacity to be effective therapeutic agents against intravascular targets.

Animals↗
PubMed1999

The effects of information framing on the practices of physicians.

OBJECTIVE: The presentation format of clinical trial results, or the "frame," may influence perceptions about the worth of a treatment. The extent and consistency of that influence are unclear. We undertook a systematic review of the published literature on the effects of information framing on the practices of physicians. DESIGN: Relevant articles were retrieved using bibliographic and electronic searches. Information was extracted from each in relation to study design, frame type, parameter assessed, assessment scale, clinical setting, intervention, results, and factors modifying the frame effect. MAIN RESULTS: Twelve articles reported randomized trials investigating the effect of framing on doctors' opinions or intended practices. Methodological shortcomings were numerous. Seven papers investigated the effect of presenting clinical trial results in terms of relative risk reduction, or absolute risk reductions or the number needing to be treated; gain/loss (positive/negative) terms were used in four papers; verbal/numeric terms in one. In simple clinical scenarios, results expressed in relative risk reduction or gain terms were viewed most positively by doctors. Factors that reduced the impact of framing included the risk of causing harm, preexisting prejudices about treatments, the type of decision, the therapeutic yield, clinical experience, and costs. No study investigated the effect of framing on actual clinical practice. CONCLUSIONS: While a framing effect may exist, particularly when results are presented in terms of proportional or absolute measures of gain or loss, it appears highly susceptible to modification, and even neutralization, by other factors that influence doctors' decision making. Its effects on actual clinical practice are unknown.

Clinical Trials as Topic↗
PubMed1999

Site-directed selection of oligonucleotide antagonists by competitive elution.

Oligonucleotide ligands that bind a protein or a small molecule of interest are readily isolated by in vitro selection and amplification of rare sequences from combinatorial libraries of sequence-randomized oligonucleotides (Gold et al., 1995). Classic systematic evolution of ligands by exponential enrichment (SELEX) protocols are affinity based (Tuerk and Gold, 1990), but because many problems and applications require antagonists, protocols for selecting inhibitors are both desirable and valuable. A widely applicable approach for isolating inhibitors is competitive elution with a molecule that binds the targeted molecule's active or binding site. We have used this approach to isolate antagonists of wheat germ agglutinin (WGA) from a library of 2'NH2-pyrimidine, 2'OH-purine oligonucleotides by elution with N N' N"-triacetylchitotriose, (GlcNAc)3. The highest affinity aptamers have equilibrium dissociation constants of 1 nM-20 nM for WGA, a 10(3)-10(4)-fold improvement relative to (GlcNAc)3, and unlike the carbohydrate, are highly specific. In addition to competing for binding with (GlcNAc)3, aptamers inhibit WGA-mediated agglutination of sheep erythrocytes, demonstrating that they are able to compete with natural ligands presented on the surfaces of cells. These results illustrate the feasibility of isolating high-affinity, high-specificity antagonists by competitive elution with low molecular weight, relatively low-affinity, and low-specificity small molecules.

Animals↗
PubMed1999

Dopamine induces neutrophil apoptosis through a dopamine D-1 receptor-independent mechanism.

BACKGROUND: For the normal resolution of an acute inflammatory response, neutrophil (PMN) apoptosis is essential to maintain immune homeostasis and to limit inappropriate host tissue damage. A delay in PMN apoptosis has been implicated in the pathogenesis of the systemic inflammatory response syndrome (SIRS). Dopamine, a biogenic amine with known cardiovascular and neurotransmitter properties, is used in patients with SIRS to maintain hemodynamic stability. We sought to determine whether dopamine may also have immunoregulatory properties capable of influencing PMN apoptosis, function, and activation state in patients with SIRS. METHODS: PMNs were isolated from healthy volunteers and patients with SIRS and treated with varying doses of dopamine and a dopamine D-1 receptor agonist, fenoldopam. PMN apoptosis was assessed every 6 hours with use of propidium iodide DNA staining and PMN function was assessed with use of respiratory burst activity, phagocytosis ability, and CD11a, CD11b, and CD18 receptor expression as functional markers. RESULTS: There was a significant delay in PMN apotosis in patients with SIRS compared with controls. Treatment of isolated PMNs from both healthy controls and patients with SIRS with 10 and 100 mumol/L dopamine induced apoptosis. PMN ingestive and cytocidal capacity were both decreased in patients with SIRS compared with controls. Treatment with dopamine significantly increased phagocytic function. Fenoldopam did not induce PMN apoptosis. CONCLUSION: Our data demonstrate for the first time that dopamine induces PMN apoptosis and modulates PMN function both in healthy controls and in patients with SIRS. These results indicate that dopamine may be beneficial during SIRS through a nonhemodynamic PMN-dependent proapoptotic mechanism.

Adult↗
PubMed1998

The binding of calcium to the B-repeat segment of SdrD, a cell surface protein of Staphylococcus aureus.

In the Sdr family of Staphylococcus aureus cell surface proteins, three recently cloned members (Josefsson, E., McCrea, K., Ni Eidhin, D., O'Connell, D., Cox, J. A., Hook, M., and Foster, T. (1998) Microbiology, in press) display variable numbers of B-repeats, i.e. segments of 110-113 residues that probably make up one folding unit. Each B-repeat contains one conserved EF-hand motif and two acidic stretches. Equilibrium dialysis revealed that segment B1-B5 of SrdD contains 14 Ca2+-binding sites with high affinity ([Ca2+]0.5, 4 microM), whereas flow dialysis yielded 5 sites of high affinity (class I) and 10 of low affinity (class II). The discrepancy could be explained by the slow induction of high affinity in the class II sites. Kinetic experiments using fluorescent Ca2+ indicators corroborated slow binding of Ca2+ at the latter sites. Circular dichroism and Trp fluorescence showed that, whereas the Ca2+ form is well folded, the metal-free form seems strongly disorganized. The Ca2+-induced conformational changes comprise both fast and slow steps, giving thus a structural support for the induction of class II Ca2+-binding sites. The B-repeats may act as rulers or springs that modulate the distance between the interactive A region and the bacterial cell surface.

Bacterial Proteins↗