Cost-effectiveness of the SQ HDM SLIT-tablet for the treatment of allergic asthma in three Eastern European Countries.

Summary: Background.The SQ® house dust mite (HDM) sublingual immunotherapy (SLIT)-tablet (ACARIZAX®, ALK-Abelló A/S, Hørsholm, Denmark) is an allergy immunotherapy tablet for people with allergic respiratory disease. This analysis aims to assess the cost-effectiveness of the SQ HDM SLIT-tablet from the perspective of three Eastern European countries: Czech Republic, Poland and Slovakia. Methods.A cost-utility model per country was developed, which compared the SQ HDM SLIT-tablet as add-on to pharmacotherapy with pharmacotherapy alone in patients with HDM allergic asthma (AA) over a five year time horizon. The effectiveness of the two interventions was based on the results from a large-scale randomised controlled trial. In the models, annual costs and quality-adjusted life year (QALY) scores from the trial were extrapolated over a five year period, and the incremental cost-effectiveness ratios (ICERs) were estimated. One-way deterministic sensitivity and scenario analyses were undertaken. Results.The SQ HDM SLIT-tablet is cost-effective in all three markets over the five year time horizon (ICERs of less than € 10,000 per additional QALY). Treatment with the SQ HDM SLIT-tablet improves patient outcomes, with QALY gains of 0.35, versus pharmacotherapy only. In all three countries, the SQ HDM SLIT-tablet also incurs increased costs compared to pharma-cotherapy treatment only. The sensitivity analysis identified utility values from the clinical trial as the main driver of the model results. Conclusion.The SQ HDM SLIT-tablet is a cost-effective treatment option for people with HDM AA in three different health care settings in Eastern Europe.

The clinical utility of PET/CT scan and tissue biopsy in the management and follow-up of paediatric Hodgkin lymphoma in South Africa.

BACKGROUND: In low- to middle-income countries (LMICs) like South Africa, there is a need to understand the clinical practices surrounding diagnosis and surveillance of paediatric Hodgkin lymphoma (HL) to reduce the burden on health systems. Understanding the clinical utility of PET/CT scans may decrease repeated tissue biopsies during disease surveillance. METHODS: This is a retrospective cohort study of patients aged less than 18 years treated for HL at Chris Hani Baragwanath Academic Hospital from 1 January 2009 to 31 December 2018. RESULTS: Fifty-four patients were included in the study; male-to-female ratio was 5:1 with a mean age of 9 years. Seventy per cent of patients (n = 38) received a PET/CT and tissue biopsy during their initial diagnostic workup, whereas 20.4% (n = 11) of patients received a PET/CT and tissue biopsy during surveillance. Tissue biopsy and PET/CT showed slight agreement (κ = 0.14) in diagnosing relapsed disease during surveillance. The false negative rate for tissue biopsy during surveillance was 42.9%. Surveillance PET/CT showed a positive predictive value (PPV) of 66.7% and negative predictive value (NPV) of 100% when compared to tissue biopsy. CONCLUSION: This study is the first cohort to explore the clinical utility of PET/CT scans and tissue biopsies in a lowresourced setting. Our findings showed slight agreement between the modalities in diagnosing relapsed disease during surveillance. A portion of this discordance can be attributed to false negative tissue biopsy results. While the sample is limited, our findings are consistent with the high NPV of PET/CT scans of > 95% as is reported in the literature.

Pelvic and acetabular fracture care in England: current workload and future directions.

INTRODUCTION: Fractures of the pelvis and acetabulum (PAFs) are challenging injuries, requiring specialist surgical input. Since implementation of the major trauma network in England in 2012, little has been published regarding the available services, workforce organisation and burden of PAF workload. The aim of this study was to assess the recent trends in volume of PAF workload, evaluate the provision of specialist care, and identify variation in available resources, staffing and training opportunity. METHODS: Data on PAF volume, operative caseload, route of admission and time to surgery were requested from the Trauma Audit and Research Network. In order to evaluate current workforce provision and services, an online survey was distributed to individuals known to provide PAF care at each of the 22 major trauma centres (MTCs). RESULTS: From 2013 to 2019, 23,823 patients with PAF were admitted to MTCs in England, of whom 12,480 (52%) underwent operative intervention. On average, there are 3,971 MTC PAF admissions and 2,080 operative fixations each year. There has been an increase in admissions and cases treated operatively since 2013. Three-quarters (78%) of patients present directly to the MTC while 22% are referred from regional trauma units. Annually, there are on average 37 operatively managed PAF injuries per million population. Notwithstanding regional differences in case volume, the average number of annual PAF operative cases per surgeon in England is 30. There is significant variation in frequency of surgeon availability. There is also variation in rota organisation regarding consistent specialist surgeon availability. CONCLUSIONS: This article describes the provision of PAF services since the reorganisation of trauma services in England. Future service development should take into account the current distribution of activity, future trends for increased volume and casemix, and the need for a PAF registry.

COT1, a gene involved in cobalt accumulation in Saccharomyces cerevisiae.

The COT1 gene of Saccharomyces cerevisiae has been isolated as a dosage-dependent suppressor of cobalt toxicity. Overexpression of the COT1 gene confers increased tolerance to cobalt and rhodium ions but not other divalent cations. Strains containing null alleles of COT1 are viable yet more sensitive to cobalt than are wild-type strains. Transcription of COT1 responds minimally to the extracellular cobalt concentration. Addition of cobalt ions to growth media results in a twofold increase in COT1 mRNA abundance. The gene encodes a 48-kDa protein which is found in mitochondrial membrane fractions of cells. The protein contains six possible membrane-spanning domains and several potential metal-binding amino acid residues. The COT1 protein shares 60% identity with the ZRC1 gene product, which confers resistance to zinc and cadmium ions. Cobalt transport studies indicate that the COT1 product is involved in the uptake of cobalt ions yet is not solely responsible for it. The increased tolerance of strains containing multiple copies of the COT1 gene is probably due to increased compartmentalization or sequestration of the ion within mitochondria.

Sacral fracture associated with small bowel entrapment: a case report.

We report the third documented case of small bowel entrapment within a sacral fracture leading to small bowel obstruction. This important diagnosis is rare and difficult to make, even with current imaging methods. We report a case in which a segment of small bowel trapped in a Denis II fracture of the sacrum required laparotomy, small bowel resection, and an omental patch over the fracture site. In this case the outcome was favorable with no residual sequelae.

Determinants and clinical impact of pressure drift in manoscan anorectal high resolution manometry system.

BACKGROUND: Pressure drift (PD), resulting from differences between room and body temperature, reduces the accuracy of pressure measurements with the Manoscan high resolution manometry (HRM) system. Our aims were to assess PD during anorectal HRM. METHODS: Defined as the residual pressure measured immediately after the catheter was removed, PD was calculated for each sensor and averaged across all 12 sensors in 454 anorectal consecutive studies recorded with 3 HRM catheters. The relationship between PD and study duration, number of prior uses of a catheter, and peak and average pressure exposure during a study were evaluated. The correction of PD with a software algorithm (thermal compensation) was evaluated in 76 studies where the most distal sensor was outside the body. KEY RESULTS: The PD varied among sensors and across catheters. The average PD (7.3 ± 0.2 mmHg) was significantly greater for newer catheters, during longer studies, or when sensors were exposed to higher pressures. Together, these factors explained 81% of the variance in overall PD. After thermal compensation, the uncorrected median PD for the most distal sensor was 2.5-5 mmHg over the study duration. Correcting this changed the interpretation (e.g., as abnormal instead of normal) of at least 1 anorectal parameter in eight of 76 studies. CONCLUSIONS & INFERENCES: During anorectal HRM, PD declines with catheter use and is greater for newer catheters, when sensors are exposed to higher pressures, and for studies of longer duration. While PD is partially corrected with thermal compensation algorithms, the impact on interpretation is modest.

The active sites of molybdenum- and tungsten-containing enzymes.

Protein X-ray crystallography has revealed the structures of the active sites of several molybdenum- and tungsten-containing enzymes that catalyze formal hydroxylation and oxygen atom transfer reactions. Each molybdenum (or tungsten) atom is coordinated by one (or two) ene-dithiolate groups of a novel pterin (molybdopterin), and the active sites are further differentiated from one another by the number of terminal oxo and/or sulfido groups and by coordinated amino acid residues. These active-site structures have no precedent in the coordination chemistry of molybdenum and tungsten.

Substrate binding and sequence preference of the proteasome revealed by active-site-directed affinity probes.

BACKGROUND: The proteasome is a multicatalytic protease complex responsible for most cytosolic protein breakdown. The complex has several distinct proteolytic activities that are defined by the preference of each for the carboxyterminal (P1) amino acid residue. Although mutational studies in yeast have begun to define substrate specificities of individual catalytically active beta subunits, little is known about the principles that govern substrate hydrolysis by the proteasome. RESULTS: A series of tripeptide and tetrapeptide vinyl sulfones were used to study substrate binding and specificity of the proteasome. Removal of the aromatic amino-terminal cap of the potent tripeptide vinyl sulfone proteasome inhibitor 4-hydroxy-3-iodo-2-nitrophenyl-leucinyl-leucinyl-leucine vinyl sulfone resulted in the complete loss of binding and inhibition. Addition of a fourth amino acid (P4) to the tri-leucine core sequence fully restored inhibitory potency. 125I-labeled peptide vinyl sulfones were also used to examine inhibitor binding and to determine the correlation of subunit modification with inhibition of peptidase activity. Changing the amino acid in the P4 position resulted in dramatically different profiles of beta-subunit modification. CONCLUSIONS: The P4 position, distal to the site of hydrolysis, is important in defining substrate processing by the proteasome. We observed direct correlations between subunit modification and inhibition of distinct proteolytic activities, allowing the assignment of activities to individual beta subunits. The ability of tetrapeptides, but not tripeptide vinyl sulfones, to act as substrates for the proteasome suggests there could be a minimal length requirement for hydrolysis by the proteasome. These studies indicate that it is possible to generate inhibitors that are largely specific for individual beta subunits of the proteasome by modulation of the P4 and carboxy-terminal vinyl sulfone moieties.

Meticillin-resistant Staphylococcus aureus in the intensive care unit: its effect on outcome and risk factors for acquisition.

BACKGROUND: Meticillin-resistant Staphylococcus aureus (MRSA) is a common cause of nosocomial infection in the intensive care unit (ICU). A perception exists that ICU-acquired MRSA is associated with poor outcomes, although there are few data to support this. AIM: To determine the effect of acquiring MRSA in the ICU on 180-day mortality, and to identify risk factors associated with acquisition. METHODS: Data were collected prospectively from 2007 to 2013. Patients who remained MRSA negative throughout their ICU admission were matched with patients who acquired MRSA in terms of age, Acute Physiology and Chronic Health Evaluation II score, length of ICU stay and surgical/non-surgical status. FINDINGS: In total, 2405 patients were included in the analysis. Patients who acquired MRSA in the ICU had significantly longer ICU stays than patients who were admitted with MRSA and patients who remained MRSA negative throughout their ICU stay (P < 0.001 for both). There were no significant differences in 180-day mortality between the groups (P = 0.238). A confirmed non-MRSA infection within 48 h of ICU admission was associated with increased risk of MRSA acquisition (adjusted odds ratio 2.57, P = 0.005), and receipt of antimicrobial therapy within 48 h of ICU admission was associated with reduced risk of MRSA acquisition (adjusted odds ratio 0.38, P = 0.014). CONCLUSION: MRSA acquisition does not contribute towards mortality in critically ill patients. This raises questions regarding the cost-effectiveness of focusing infection prevention measures on the control of MRSA in ICUs. The low acquisition rate and lack of risk factors identified for MRSA in the study cohort indicate that efforts should be directed towards continual improvement of standard infection control procedures for all patients.

Experience of managing open fractures of the lower limb at a major trauma centre.

INTRODUCTION: In April 2012 the John Radcliffe Hospital in Oxford became a major trauma centre (MTC). The British Orthopaedic Association and British Association of Plastic, Reconstructive and Aesthetic Surgeons joint standards for the management of open fractures of the lower limb (BOAST 4) require system-wide changes in referral practice that may be facilitated by the MTC and its associated major trauma network. METHODS: From 2008 to 2013 a multistep audit of compliance with BOAST 4 was conducted to assess referral patterns, timing of surgery and outcomes (surgical site infection rates), to determine changes following local intervention and the establishment of the MTC. RESULTS: Over the study period, 50 patients had soft tissue cover for an open lower limb fracture and there was a significant increase in the proportion of patients receiving definitive fixation in our centre (p=0.036). The median time from injury to soft tissue cover fell from 6.0 days to 3.5 days (p=0.051) and the median time from definitive fixation to soft tissue cover fell from 5.0 days to 2.0 days (p=0.003). The deep infection rate fell from 27% to 8% (p=0.247). However, in 2013 many patients still experienced a delay of >72 hours between injury and soft tissue cover, primarily owing to a lack of capacity for providing soft tissue cover. CONCLUSIONS: Our experience may be relevant to other MTCs seeking to identify barriers to optimising the management of patients with these injuries.

Bioavailability of topically administered steroids: a "mass balance" technique.

The percutaneous absorption of four steroids (hydrocortisone, estradiol, testosterone, and progesterone) has been measured in vivo in man under occluded and "protected" (i.e., covered, but non-occlusive) conditions. The experimental approach, involving simple modifications of standard radiochemical methodology, has enabled excellent "mass balance" and dose accountability to be achieved. Consequently, the utility of the procedure for the measurement of in vivo topical bioavailability can be inferred. In addition, because of the precision and accountability of the results, the technique offers a potential means to establish quantitative structure-penetration relationships for skin absorption in man. It was found that steroid absorption increased with increasing lipophilicity up to a point, but that penetration of progesterone (the most hydophobic analog studied) did not continue the trend and was at least partly rate-limited by slow interfacial transport at the stratum corneum-viable epidermis boundary. Comparison of data obtained from the occluded and "protected" experiments permitted the effect of occlusion (defined as the complete impairment of passive transepidermal water loss at the application site) to be assessed. Occlusion significantly increased percutaneous absorption of estradiol, testosterone, and progesterone but did not effect the penetration of hydrocortisone. A mechanism is proposed to explain why the absorption of the more lipophilic steroids is enhanced by occlusion but that of the most water-soluble (i.e., hydrocortisone) is not. It is suggested that the rate-determining role of the sequential steps involved in percutaneous absorption can be revealed by experiments of the type described using related series of homologous or analogous chemicals.

Ovarian and peripheral blood inhibin concentrations increase with gonadotropin treatment in immature rats.

The effects of PMSG treatment on ovarian and circulating inhibin concentrations in immature female rats has been examined. Sixty-four hours after injection of 10, 20 or 40 IU of PMSG the animals were anesthetized with ether; ovaries, uteri and blood from the abdominal aorta were collected. Steroid-free extracts of ovary and serum samples were prepared and assayed quantitatively for inhibin activity in an in vitro bioassay system. PMSG treatment elevated (p less than 0.001) both uterine and ovarian wt, and ovarian and peripheral concentrations of inhibin. A dose-related increase occurred ovarian wt, and in peripheral and ovarian content of inhibin. Ovarian inhibin concentration increased with dose of PMSG until the highest dose, where a significant decline and luteinization were seen. Peripheral FSH levels were significantly lowered at all doses of PMSG treatment; in contrasts, LH was significantly elevated, due to cross-reaction of PMSG in the LH assay. These results show that both ovarian and circulating levels of inhibin are related to the degree of gonadotropic stimulation, supporting the view that inhibin is involved in folliculogenesis and in the feedback regulation of FSH.

Involvement of the plasmin system in dissociation of the insulin-like growth factor-binding protein complex.

A variety of treatments, including acid, heparin, and proteases, are known to free insulin-like growth factors (IGFs) from their binding proteins (IGFBPs). However, the physiologically relevant mechanism regulating the interaction of IGFs and IGFBPs is unknown. We report here the ability of plasmin to dissociate IGFs from IGFBPs. In chromatographic experiments, plasmin completely dissociated complexes of [125I] IGF-I-BP and [125I]IGF-II-BP formed with purified decidual IGFBP (hIGFBP-1) or IGFBPs present in medium conditioned by human osteosarcoma MG-63 cells. Plasmin dissociation of IGF-BP complexes was dose dependent. Neither plasminogen nor plasminogen activators (PAs) alone affected dissociation; however, activation of plasminogen to plasmin by either urokinase PA or tissue-type PA resulted in the dissociation of IGF-BP complexes. Plasmin dissociated immunoreactive and bioactive IGF from IGFBP equivalent to approximately 70% and approximately 60% of the acid control value, respectively. In medium conditioned by MG-63 cells, dissociation of IGF-BP complexes was catalyzed by PAs secreted by MG-63 cells, principally urokinase PA. Limited plasmin degradation of IGF was suggested by chromatographic experiments involving [125I] IGF. Treatment of uncomplexed IGF-I with plasmin concentrations equivalent to those in chromatographic experiments did not result in a significant loss of bioactivity, although a 2-fold increase in the plasmin concentration resulted in a approximately 20% loss of activity. Similar plasmin treatment of equimolar concentrations of hIGFBP-1 resulted in a marked degradation of IGFBP, with loss of IGF-binding ability. In vitro experiments confirmed plasmin dissociation of bioactive IGF-I from hIGFBP-1. In MG-63 cells, IGFBPs can form an IGF reservoir in the pericellular space surrounding the cells by combining IGFs with IGF-BP to form complexes that are incapable of binding to the IGF receptors. The secretion of PAs by osteosarcoma cells and the availability of plasminogen in the extravascular tissues indicate the possibility of a regulatory system in osteosarcoma cells in which pericellular plasmin affects the availability of IGFs to their membrane receptors.

Polypharmacy among patients attending an AIDS clinic: utilization of prescribed, unorthodox, and investigational treatments.

The objective of this study was to describe the utilization and characteristics associated with the use of prescribed, over-the-counter, investigational, and unorthodox treatments among AIDS clinic patients. This report is derived from cross-sectional data obtained using structured telephone surveys. Study participants (n = 197) were recruited from the University of California, San Francisco, Medical Center AIDS clinic. One hundred eighty-nine participants (96%) received 1-24 prescription medications during the 3 months prior to interview. Those with an AIDS diagnosis received a relatively greater number of prescription drugs (p = 0.0001); an average of 5.6 prescribed medications were used by AIDS patients versus 4.8 among AIDS-related complex and 2.3 among asymptomatic patients. Thirty-one percent participated in drug trials during the 3 months before interview, including 18% who were in multiple studies. Twenty-nine percent used unorthodox treatments. Seventy-five (40%) received prescription medication from a provider other than their primary provider. A more advanced stage of illness was associated with the use of unorthodox treatments (p = 0.003): users of these treatments had a greater educational attainment than nonusers (p = 0.03) and were significantly less likely to report that their primary provider was aware of all the treatments they used (odds ratio = 2.1, p less than 0.03). We conclude that use of polypharmacy among some AIDS clinic patients is common, could create an increased risk for adverse drug reactions, and may affect clinical drug trials. Despite having decided to obtain care at a university-based clinic, many of the participants of this study also chose to receive unorthodox therapies and care from nonprimary medical providers.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of steady-state follistatin mRNA levels in rat granulosa cells in vitro.

The regulation of steady-state follistatin mRNA levels by different pituitary hormones and peptide factors was examined in granulosa cell cultures derived from diethylstilboestrol-treated immature rats. Cytosolic RNA from cell cultures was prepared by lysis and equal amounts of RNA from all samples were analysed with a solution-hybridization assay using a 32P-labelled antisense probe corresponding to a part of exon 5 together with a part of the 5' end of exon 6 of the rat follistatin gene. In addition, a specific 35S-labelled probe for cyclophilin was used as an internal standard. The results show that 5 micrograms FSH/l for 24 to 72 h stimulated steady-state follistatin mRNA levels, reaching levels 18.5-fold higher than controls. LH (0.2-100 micrograms/l) had only minor effects on follistatin mRNA levels in FSH-primed granulosa cells and prolactin, GH and IGF-I did not show any significant effects. Activin raised basal as well as FSH-stimulated steady-state follistatin mRNA levels up to ten- and twofold above controls respectively, whereas epidermal growth factor was found to inhibit FSH-stimulated follistatin mRNA levels in a dose-dependent manner. It is concluded that follistatin mRNA levels in granulosa cells are regulated by FSH rather than LH, and that the stimulation by FSH can be inhibited by epidermal growth factor but enhanced by activin. Activin alone was also capable of stimulating follistatin mRNA.

Assay of cell-mediated cytotoxicity by tritiated thymidine labeled tumor cells.

A microtechnique for quantitating cell-mediated cytotoxicity using tritiated thymidine labeled target cells is presented. Target cell incorporation of tritiated thymidine is augmented by using methotrexate to arrest endogenous thymidine synthesis. Complete solubilization of labeled cells is accomplished by oxidizing samples to tritiated water and carbon dioxide prior to quantitation in a scintillation counter. This versatile technique, using various combinations of lymphocytes, sera and target cells, permits the simultaneous comparison of 500 or more individual samples.

Ovarian inhibin content in immature female rats following hypophysectomy: control by exogenous gonadotrophins.

The effect of exogenous ovine gonadotrophins on ovarian inhibin content in hypophysectomized immature female rats was studied to determine which of the pituitary gonadotrophins control ovarian inhibin production in vivo. Inhibin and steroid contents of 27,000 g ovarian supernatants were measured by specific radioimmunoassay. Whereas injection of ovine (o)FSH resulted in dose-dependent increases in ovarian weight, progesterone and oestradiol content above that of control animals injected with diethylstilbestrol alone, injection of oLH was without significant effect. Similarly, ovarian inhibin content was not altered significantly by injection of oLH, but was increased in a dose-dependent manner by injection of oFSH. Concentrations of inhibin in serum were correlated directly with ovarian inhibin content. The results of this study indicate that ovarian inhibin content is increased by injection of oFSH but not oLH. It is likely, therefore, that inhibin production by small antral follicles is controlled by FSH activity alone.

A phenoxyl radical complex of copper(II).

A new N,O-bidentate pro-ligand (HL), [ML2] (M = Cu, Zn) and [CuL2][BF4] have been synthesised; [CuL2].4DMF and [CuL2][BF4].2CH2Cl2 have been crystallographically and spectroscopically characterised; these data indicate that [CuL2]+ cations are constituted as [Cu2+(L.)(L-)]+ and involve the phenoxyl radical L..

Acquisition of resistance to 6-azauridine through DNA amplification in neoplastic but not normal osteoblasts.

In this communication, we have characterized the resistance to AZUrd in tumorigenic mouse C3H-OS osteosarcoma cells and non-tumorigenic MC3T3-E1 osteoblast cells. DNA and RNA blot analysis showed a 30-fold increase in UMP synthase specific DNA and a 10-fold increase in mRNA, respectively, in resistant versus non-resistant C3H-OS cells. No corresponding increases in either UMP synthetase DNA or mRNA were evident in resistant MC3T3-E1 osteoblasts. Karyotype analysis of MC3T3-E1 and C3H-OS cells revealed translocations in the resistant cells. Regardless of drug-sensitive or resistant phenotype, the normal and neoplastic cells exhibited aneuploidy which was significantly more pronounced in the non-resistant tumor cells. Additionally, the number of chromosomes decreased in all resistant cells whether normal or neoplastic. We conclude that genomic instability in neoplastic cells is a prerequisite for the generation of drug resistant variants via the process of gene amplification.

An MCD spectroscopic study of the molybdenum active site in sulfite oxidase: insight into the role of coordinated cysteine.

Temperature-dependent magnetic circular dichroism (MCD) spectroscopy has been used for the first time to probe the electronic structure of the Mo active site in sulfite oxidase (SO). The enzyme was poised in the catalytically relevant [Mo(V):Fe(II)] state by anaerobic reduction of the enzyme with the natural substrate, sulfite, in the absence of the physiological oxidant cytochrome c. The [Mo(V):Fe(II)] state is of particular importance, as it is proposed to be a catalytic intermediate in the oxidative half reaction, where SO is reoxidized to the resting [Mo(VI):Fe(III)] state by two sequential one-electron transfers to cytochrome c. The MCD spectrum of the enzyme shows no charge transfer transitions below approximately 17000 cm(-1). This has been interpreted to result from (1) a severe reduction in ene-1,2-dithiolate sulfur in-plane and out-of-plane p orbital mixing, (2) a decrease in the dithiolate sulfur out-of-plane p-Mo d(xy) orbital overlap, and (3) an orthogonal orientation between the vertical cysteine sulfur p (perpendicular to the Mo-Scys sigma-bond) and Mo d(xy) orbitals. The spectroscopically determined cysteine sulfur p-Mo d(xy) bonding scheme in the [Mo(V):Fe(II)] state is consistent with the crystallographically determined O-Mo-Scys-C dihedral angle of approximately 90 degrees and precludes a covalent interaction between the vertical cysteine sulfur p orbital and Mo d(xy), effectively decoupling the cysteine from an effective through-bond electron transfer pathway. We have tentatively assigned a 22250 cm(-1) positive C-term feature in the MCD as the cysteine S(sigma)-->Mo d(xy) charge transfer that becomes allowed by a combination of configuration interaction and low-symmetry; however, the orbital overlap is anticipated to be quite small due to the near orthogonality of these orbitals. Therefore, we propose that the primary role of the coordinated cysteine is to decrease the effective nuclear charge on Mo by charge donation to the metal, statically poising the active site at more negative reduction potentials during electron transfer (ET) regeneration. Finally, the results of this study are consistent with the pyranopterin ene-1,2-dithiolate acting to couple the Mo site into efficient superexchange pathways for ET regeneration following oxygen atom transfer to the substrate.