monocytogenes to β-lactams selleck products was examined. Deletion mutant in lmo1941 was constructed and subjected to studies, which revealed that the deletion of lmo1941 had no effect on susceptibility and tolerance to penicillin G and ampicillin but resulted, however, in increased susceptibility of L. monocytogenes to several cephalosporins. Subsequently, the potential effect of lmo1941 mutation on the cell wall of L. monocytogenes was investigated. The analysis revealed quantitative changes in the muropeptide profile of peptidoglycan and a decrease in density of the high-density zone of cell wall of the mutant strain. Both these changes were observed in cells taken from the stationary phase. These results indicate that the surface protein

Lmo1941 affects peptidoglycan composition and cell wall structure of L. monocytogenes in the stationary phase of growth. “
“The genus Rickettsiella comprises intracellular bacterial pathogens of a wide range of arthropods

that are currently classified in four recognized species and numerous further pathotypes. However, both the delineation of and the synonymization of pathotypes with species TSA HDAC supplier are highly problematic. In the sequel of a previous phylogenomic study at the supra-generic level, nine selected genes – the 16S and 23S rRNA genes and the protein-encoding genes dnaG, ftsY, gidA, ksgA, rpoB, rpsA, and sucB – were evaluated for their potential as markers for the generic and infra-generic taxonomic classification of Rickettsiella-like bacteria. A methodological approach combining phylogenetic reconstruction with likelihood-based significance (-)-p-Bromotetramisole Oxalate testing was employed on the basis of sequence data from the species Rickettsiella grylli and Rickettsiella popilliae, pathotypes ‘Rickettsiella melolonthae’ and ‘Rickettsiella tipulae’. This study provides the first multilocus sequence typing (MLST) data for the genus Rickettsiella and identifies two new genetic markers, gidA and sucB, for the infra-generic classification within this taxon. In particular, aforesaid genes were found more reliable

and informative markers than the corresponding 16S rRNA-encoding sequences that failed to produce strictly significant infra-generic taxonomic assignments. However, gidA- and sucB-based phylogenies were consistent with the currently accepted view of species delineation and species-pathotype synonymization within the genus Rickettsiella. The genus Rickettsiella (Philip) comprises intracellular bacterial pathogens of a wide range of arthropods. The currently valid taxonomy of these bacteria (Garrity et al., 2005) is primarily based on the indication of a strain’s original host. Moreover, the resulting pathotype designation is partially superposed by the morpho- and serologically founded distinction of four recognized species, namely the nomenclatural type species Rickettsiella popilliae (Dutky & Gooden), Rickettsiella grylli (Vago & Martoja), Rickettsiella chironomi (Weiser), and Rickettsiella stethorae (Hall & Badgley).

, 1997). In the symbiosis of trypanosomatids, intense metabolic exchanges occur between both partners: the bacterium contains essential enzymes that complete important biosynthetic pathways of the protozoa and in exchange receives suitable physical conditions and energy supply from the host (reviewed by Motta, 2010). As in most prokaryotes, sterols are absent in symbiont membranes that have cardiolipin (CL) as the major phospholipid, followed by similar amounts of PC and PE and a minor quantity of PI (Palmié-Peixoto et al., 2006). The endosymbiont enhances the protozoan phospholipid production and depends in part on its host cell to obtain PC (Azevedo-Martins et al.,

2007). Organelles of symbiotic origin play Akt tumor important roles in the eukaryotic cell lipid biosynthesis. Significant levels of phospholipid production occur in mitochondria that synthesize phosphatidic acid (PA) and phosphatidylglycerol, which is used to produce CL, a lipid that is mainly found in prokaryotes and mitochondria, as well as PE (Van Meer et al., 2008). In this work, we tested the effect of miltefosine RG7422 ic50 on cell proliferation, ultrastructure, and phospholipid biosynthesis of A. deanei. The main proposal of miltefosine treatment on this nonpathogenic trypanosomatid species is to evaluate, if once the protozoan

phospholipid production is affected, how does it influence the symbiotic bacterium and mitochondrion composition. Thus, it is worth considering that both structures have symbiotic origin and are related to the protozoan phospholipid metabolism. Angomonas deanei was grown Exoribonuclease at 28 °C for 24 h in Warren’s culture medium (Warren, 1960) supplemented with 10% fetal calf serum. Miltefosine (Cayman Chemical) was used after dilutions of a 100 mM stock solution dissolved in absolute methanol. Cells (1.0 × 106 mL−1) were inoculated in culture medium and after 12 h (exponential growth phase) were submitted to different drug concentrations: 10, 25, 50, 75, and 100 μM. Protozoa were collected from the culture at 12 h intervals until 72 h of growth;

then, part of the cells were counted in a Neubauer chamber, and the remainder was processed for transmission electron microscopy as described below. To verify the effect of miltefosine on phospholipid biosynthesis, cells were grown for 24, 36, and 48 h in Warren medium containing 4 μCi of 32Pi, whereas for cell fractioning assays, protozoa were cultivated for 24 h in the presence of 10 μCi of 32Pi. Isolated symbionts and mitochondria were obtained by cell fractioning as established by Alfieri & Camargo (1978), with some modifications. Cells were disrupted using an ultrasonic disruptor GEX-600 (three series of 15-s pulses at 10% amplitude). The homogenate passed throws differential centrifugations and sucrose gradient, to obtain a rich fraction of endosymbionts and mitochondria. Then, fractions were resuspended in 1 mL of Tris-HCl 20 mM and sucrose 0.

05, P = 3.9 × 10−4) and SCN-lesioned (effect of brain area, F3,61 = 2.50, P = 0.068) rats, and they did not differ between the R-MAP and R-Water groups in either SCN-intact rats (interaction between brain area and treatment, F3,60 = 0.91, P = 0.44; main effect of treatment, F1,60 = 3.3 × 10−4, P = 0.99) or SCN-lesioned rats (interaction Dactolisib mw between brain area and treatment, F2,46 = 0.22, P = 0.81; main effect of treatment, F1,46 = 0.21, P = 0.65 for SCN-lesion; Fig. 8B). When compared between the SCN-intact and SCN-lesioned rats, the damping rates did

not differ in either the R-MAP group (interaction between brain area and SCN-lesion, F2,46 = 0.22, P = 0.81; main effect of SCN-lesion, F1,46 = 0.21, P = 0.65) or the R-Water group (interaction between brain area and SCN-lesion, F3,55

= 1.92, P = 0.14; main effect of check details SCN-lesion, F1,55 = 0.95, P = 0.33). The numbers of slices examined were as follows: (i) in the SCN-intact rats: SCN, R-Water, 9; R-MAP, 9; OB, R-Water, 9; R-MAP, 9; CPU, R-Water, 7; R-MAP, 8; PC, R-Water, 5; R-MAP, 1; and SN, R-Water, 9; R-MAP, 8, and (ii) in the SCN-lesioned rats: OB, R-Water, 9; R-MAP, 10; CPU, R-Water, 8; R-MAP, 9; PC, R-Water, 8; R-MAP, 8; and SN, R-Water, 9; R-MAP, 8. The present study clearly demonstrates that restricted MAP drinking at a restricted time of day not only induced MAO in behavior but also entrained it. The free-running of MAO under ad-MAP was modified by the SCN circadian pacemaker entraining to LD. MAO was also expressed in the circadian Per2 rhythms in several extra-SCN brain areas. The Per2 rhythms were phase-shifted by R-MAP. The phase shifts were accelerated by the SCN lesion, especially in the OB and SN, indicating dual regulation of the extra-SCN circadian oscillators in the brain by the SCN and MAO. In the absence of the SCN circadian pacemaker, R-Water also induced circadian oscillation which was not identical with MAO. The oscillatory mechanism underlying MAP-induced behavioral rhythm (i.e., MAO) is suggested as consisting of several extra-SCN oscillators in the brain (Masubuchi et al., 2000) but the exact mechanism is not well understood. A Edoxaban success of ex

vivo analysis of MAO (Natsubori et al., 2013a,b) opened a new experimental approach to this issue, and the fixation of the MAO phase by R-MAP in the present study enabled us to analyse the phase relationships among extra-SCN oscillators in the brain more precisely. The induction of MAO by R-MAP was revealed by subsequent ad-MAP, where the enhanced behavior components at the time of restricted MAP supply showed phase-delay shifts with a period > 24 h. Acceleration and deceleration of phase-delay shifts in MAP-induced behavioral rhythm were observed in the SCN-intact rats but not in the rats with bilateral SCN lesions (Figs 1 and 2). The rate of phase-delay shifts in the SCN-lesioned rats was 1.3 h/day on average and corresponded to a free-running period of 25.3 h.

Conidiophores arising from submerged hyphae 4–6 μm in length, occasionally forming loose synnemata up to 2 mm high; stalks with roughened thick walls 3–4 μm wide consisting of verticillate branches with whorls of two to four phialides. Phialides 6–9 × 2.5–3 μm, having a swollen basal portion tapering into a short distinct neck about 1 μm wide. Conidia in divergent chains,

ellipsoidal to fusiform, smooth-walled to slightly roughened, hyaline, purple en masse, 2–3 × 2–4 μm. Conidial structures formed near the agar atypical: phialides solitary or in verticils, 2–4, variable in length (Fig. 3g and h); shaped like typical Purpureocillium lilacinum phialides, or very long (up to 30 μm) and Acremonium-like. Cylindrical, occasionally slightly curved conidia formed in ‘slimy heads’ on these Acremonium-like structures, conidia on these structures variable Dabrafenib datasheet in size, measuring 2.0–14 × CH5424802 manufacturer 1.5–2.5 μm

(Fig. 3i). This conidiogenesis was also observed by Okada et al. (1995) for P. nostocoides (=Purpureocillium lilacinum). Chlamydospores absent. Species previously assigned to Paecilomyces causing human mycoses include Paecilomyces farinosus, Paecilomyces javanicus, P. lilacinus, P. marquandii, Paecilomyces taitungiacus (=anamorph of Thermoascus taitungiacus), P. variotii and Paecilomyces viridis. Of these, P. variotii is retained in the genus Paecilomyces (as it is the type), P. javanicus and P. farinosus CHIR-99021 have been returned to

the genus Isaria in the Hypocreales (Luangsa-ard et al., 2004), P. viridis has been transferred to Chamaeleomyces (Sigler et al., 2010) and P. lilacinus is accommodated here in the genus Purpureocillium. P. marquandii is currently maintained in Paecilomyces; however, this species is unrelated to P. variotii and should to be transferred to a new genus. Paranomuraea was suggested for P. marquandii and Paecilomyces carneus (Domsch et al., 2007), but this genus has yet not been published validly. Samson (1974) considered P. lilacinus and P. marquandii to be very close to each other, based on overall morphology and spore color. Paecilomyces marquandii differs from Purpureocillium lilacinum by its hyaline conidiophores and the typical yellow reverse. Although both species have a similar morphology, phylogenies show them to be separated in two families of the Hypocreales (Sung et al., 2007). Some clinical isolates have been identified as P. marquandii (Castro et al., 1990; Naldi et al., 2000). These isolates need to be re-examined using sequence-based methods to determine whether P. marquandii genuinely has the potential for human pathogenicity or whether this is merely a misidentification of Purpureocillium lilacinum. Correct identification is crucial because Purpureocillium lilacinum is significantly more resistant to amphotericin B than P. marquandii (Aguilar et al., 1998).

, 2013). In the current study, PD0325901 solubility dmso these fixational eye movements are more pronounced in ASD, due to slow drifts, as no significant differences in estimates of microsaccades were found between groups. It is extremely unlikely that the small differences (< 2 min of arc) in the standard deviation of eye position could account for the differences in amplitude of visual evoked responses observed here. Recall that the stimuli used herein subtended fully 6° of visual space, and this variance between groups is two orders of magnitude smaller. Further, the same levels of eye-position variability were also observed

for centrally presented stimuli where we found no differences in evoked response between groups. In recent years a number of studies have provided evidence for common generators of saccades and microsaccades (Martinez-Conde et al., 2013). The observed higher variability of eye position during fixation in the ASD participants might therefore

mirror studies that reported problems in oculomotor control for saccadic eye movements (Goldberg et al., 2002; Takarae et al., 2004; Stanley-Cary et al., 2011), with higher variability of landing positions in ASD participants. While an altered cortical representation account of the present findings is the most parsimonious with existing evidence in our view, there are other explanations that should be considered. When humans deploy attention covertly, stimuli JQ1 at the attended

Tau-protein kinase peripheral location receive enhanced processing. This commonly results in greater P1 amplitudes in VEP (Hillyard & Anllo-Vento, 1998; Kelly et al., 2008) and VESPA studies (Frey et al., 2010). Therefore, one possibility is that ASD participants covertly attended the peripherally presented stimuli, or maintained a broader focus of attention than their neurotypical peers. A purely attentional account, however, seems unlikely. We employed an attention-demanding central task during peripheral stimulation. Because neither behavioral performance nor eye-tracking measures differed between ASD and TD groups, it seems likely that both deployed attention equally to the central fixation task. Cortical remapping could account for a number of reported results on visual functioning in autism. For example, it has been reported that individuals with autism exhibit superior performance in visual search tasks (Plaisted et al., 1998; O’Riordan et al., 2001). Such a pattern of results could be explained by an enhanced representation of peripheral space, allowing individuals with ASD to explore their visual environment more efficiently. Another finding that could potentially relate to enhanced representation of peripheral space is lateral glance behavior (Mottron et al., 2007), frequently exhibited by a subpopulation of children with ASD.

Half (53.4%) of respondents attended postpartum diabetes screening. Barriers to screening included a lack of awareness of the need to attend screening, the inconvenience associated with the two to three hour length of the OGTT, and the need to attend screening with infants and young children. Reported facilitators included improved awareness of the need for screening, multiple reminders, and a more pleasant and convenient test. Facilitation strategies aimed at increasing the

awareness of postpartum diabetes risks and promoting the provision of accurate and consistent screening advice from medical providers may assist in improving attendance at postpartum diabetes screening. Tyrosine Kinase Inhibitor Library A more acceptable screening test and establishment of a national database for routine screening reminders may also encourage CT99021 chemical structure women to attend postpartum diabetes screening. Copyright © 2011 John Wiley & Sons. “
“During pregnancy, the term diabetic nephropathy is used to describe an heterogenous group of patients with either microalbuminuria or overt nephropathy (various degrees of proteinuria) with or without maternal hypertension or significant impairment in renal function, and often associated with diabetic

retinal microvascular disease. During the past decade, several studies have reported pregnancy outcomes in patients with various stages of diabetic nephropathy. The overall perinatal survival rate was 95%; however, these pregnancies Megestrol Acetate continue to be associated with very high rates of superimposed pre-eclampsia (32–65%) preterm delivery (57–91%), and fetal growth restriction (12–45%). Comprehensive evaluation prior to conception or early in pregnancy will permit appropriate counseling and allow for the implementation of targeted strategies to improve pregnancy outcome. There is a general agreement that tight

control of blood glucose and blood pressure prior to conception and throughout gestation, in association with frequent monitoring of maternal and fetal wellbeing along with timely delivery, are the key elements to improved pregnancy outcome. Drugs acting on the renin angiotensin system should be discontinued at conception. The majority of reported studies suggest that pregnancy per se does not increase the risk of progression to end-stage renal disease in patients with mild renal impairment prior to conception. “
“Young patients with diabetes are particularly vulnerable to long-term complications, and require a carefully planned transition to adult diabetes care. As clinic non-attendance has been identified as an issue for transitional clinics, we audited our well established clinic to look at non-attendance rates, and to examine the characteristics of those who miss transitional clinic appointments. We conducted a retrospective analysis of audit data from the diabetes transitional clinic in January to December 2004, and September 2007 to September 2008.

Our study focused primarily on the suitability of single active ingredient analgesics; however, a number of fixed-dose combination selleck analgesics are available in the OTC setting. From a suitability perspective their

use requires even more care, making it important to ensure that consumers are aware of the potential risks associated with both active ingredients when selecting these products. Our research found no significant public health issues associated with the OTC use of paracetamol, but it has shown that up to three in 10 regular users of OTC NSAIDs have current or prior medical conditions that warrant discussion with a healthcare professional prior to their use. It is important to note that

some of these consumers may already be acting upon such advice, reducing the potential risk. However, with a large proportion of regular users of OTC NSAIDs purchasing these products outside the pharmacy setting, the quality use of OTC NSAIDs is becoming increasing reliant on product labelling and the ability of consumers to understand and self-assess their own level of risk. A key theme emanating from our data and from other recent changes in the analgesics landscape both locally and globally is the continued need to ensure a high level of consumer education ABT-737 price regarding the appropriate choice and use of analgesics. For the vast majority of consumers who have used these medications in the past the potential risks are minimal. However, consumers need to be aware that if their health status changes then this warrants a discussion with a healthcare professional to confirm the continued appropriateness of their OTC analgesic medication. Rather than placing the onus solely on the consumer to actively seek advice and hoping that this is undertaken a more practical approach would be to also reinforce with healthcare professionals

the need to proactively probe patients about the use of OTC analgesics and offer advice as to any changes that need be undertaken when they present with a new condition that puts them into an at-risk population. The safe and Mannose-binding protein-associated serine protease effective use of any OTC medication requires active participation and open communication between the user and healthcare professionals. Our study demonstrates that since ibuprofen has become available outside the pharmacy setting in Australia fewer people are using NSAIDs appropriately according to the label; compared to 2001, in 2009 10.2% more regular OTC analgesic users were using ibuprofen despite having contraindications, warnings, precautions or potential drug-interactions. The increasing use and wider availability of OTC NSAIDs may have led to a more relaxed attitude regarding the use of these medicines.

Size exclusion chromatography was performed using a Superdex 75 column (10 × 16 mm, 22 mL bed volume)

equilibrated in 100 mM HEPES buffer, pH 7.5, containing 150 mM NaCl at 0.5 mL min−1. The apparent relative molecular mass of the native protein was determined by comparing its retention time (monitored PD0325901 molecular weight at 280 nm) with that of standard proteins (albumin 67 kDa, ovalbumin 43 kDa, chymotrysinogen A 25 kDa, RNAse A 13.7 kDa). EMSA (Kerr, 1995) was performed in a 20-μL assay mixture consisting of 20 mM HEPES, 20 mM KCl, 5 mM MgCl2, 2 mM DTT, 10% v/v glycerol, 0.5 mg mL−1 BSA, pH 8.0, 800 ng competitor DNA (chromosomal DNA of E. coli Rosetta), 1-pmol DNA fragment of interest and 1–10 pmol purified AtuR. Binding was allowed for a period of 20 min at room temperature, after which aliquots of the assay mixture were mixed with loading solution and were run on a 5–12% w/v polyacrylamide gel at 5 °C. After electrophoresis, the gel was stained with ethidium bromide. Staining with ethidium bromide turned out to be sufficient to visualize gel mobility shifts. DNA concentrations were determined using the NanoDrop system. Expression of the atu gene cluster is strictly regulated in P. aeruginosa and is repressed during growth on unrelated substrates such as nutrient broth, glucose or succinate. Accordingly, no geranyl-CoA carboxylase (GCase), the key enzyme of the Atu pathway, Belinostat can be detected

in cell extracts of glucose or succinate cells (Hector & Fall, 1976; Fall & Hector, 1977; Fall, 1981; Höschle et al., 2005; Aguilar et al., 2008). When P. aeruginosa was cultivated in the presence of isovalerate Wilson disease protein (or leucine), the genes of the leucine and isovalerate utilization (Liu) pathway are induced as revealed by 2-D gel electrophoresis and by detection of methyl-crotonyl-CoA carboxylase (MCase) protein in cell extracts (Fig. 1a) (Förster-Fromme et al., 2006).

MCase is the key enzyme of the Liu pathway. However, GCase or other Atu proteins are not detectable in isovalerate-grown cells. The expression of the GCase (subunits AtuC and AtuF) and of the other atu gene cluster products requires the presence of acyclic monoterpenes such as citronellol, geraniol or the respective aldehydes (citronellal, geranial) or acids (citronellate, geranylate) during growth of the bacteria (Fig. 1a) (Förster-Fromme et al., 2006). Growth on compounds with the same number of carbon atoms in the backbone as monoterpenes, but without branched methyl groups such as octanol or octanate, did not result in the formation of MCase or GCase (Fig. 1a). Citronellol-grown cells also expressed the proteins of the Liu pathway apparently because the end product of the Atu pathway and subsequent β-oxidation, methyl-crotonyl-CoA, enters the Liu pathway (Fig. S1, Fig. 1, lane ‘Cs’). In conclusion, the expression of atu genes is highly regulated.

Saddle and nasolabial angles are significantly greater in RDEB than normal50. The changes in facial skeleton may reflect reduced nutritional intake PI3K Inhibitor Library (feeding problems) and subsequent reduced bone growth50. Additionally, or alternatively, perioral soft tissue scarring during early childhood may result in reduced size of the jaws84. Bone atrophy/osteoporosis.  Osteoporosis has been increasingly identified in patients with this form of RDEB in recent years56. Radiographic records and computerized tomography scans of the jaw revealed extensive bone atrophy of the jaws in six of six patients31. During surgery, the alveolar ridges of these patients were found to be atrophic

in all cases23,31. Kindler syndrome has only recently been added as part of the classification of EB58. Only few case reports of patients with Kindler syndrome describe their oral features34,85–90. The evidence suggests that patients with Kindler syndrome can present with fragile mucosa, microstomia, and partial vestibule obliteration, although microstomia was not identified in all patients with Kindler syndrome34,85,86. Special attention has been given Trametinib ic50 to periodontal disease, which was initially reported in two patients34,88. Thereafter, a series

of 18 patients was compared to healthy controls, revealing that patients with Kindler syndrome have a higher prevalence (72%vs 46%), earlier onset, and faster progression of periodontitis85.

Squamous cell carcinoma of the hard palate has also been reported in a patient with this condition86. Inherited epidermolysis bullosa (EB) comprises a group of genetically and clinically heterogeneous diseases characterized by the formation of blisters and erosions on skin and mucous membranes following minor traction or trauma26. It is caused by mutations in the genes encoding proteins of the dermal–epidermal Dolutegravir supplier adhesion zone91. 7.3.1 Classification of EB.  EB presents a wide range of clinical phenotypes with over 1000 mutations identified in 13 structural genes. Classification schemes were first introduced by Pearson in 196292. Since then, various consensus classifications have been published58,93,94. The current classification scheme begins with the separation of EB into four major types based on the level of blister formation into EB simplex (EBS, intra-epidermal), junctional EB (JEB), dystrophic EB (DEB, dermolytic), and Kindler syndrome (mixed levels). Patients are then separated by major and minor EB subtypes. The expanded classification scheme includes the following: four types, seven major subtypes, and 33 minor subtypes58. A summary of this classification system is presented in Table 1. 7.3.2 General clinical manifestations.  The hallmark feature of inherited EB is mechanical fragility of the skin and the appearance of vesicles and bullae36.

Protein content in biological samples was determined

using the Coomassie Blue dye-binding procedure of Bradford (1976). Proteins were separated in 7.5% SDS-PAGE (Gallagher et al., 1992), and the resolved proteins were stained with Coomassie Blue R250. Recombinant wild type and mutant apoforms of LH were activated on the addition of 4 mM CaCl2 and 200 μM PQQ followed by subsequent incubation at room temperature for 1 h. The LH activity was measured spectrophotometrically using horse heart cytochrome c as the electron acceptor (Hopper et al., 1991). A unit of LH enzyme activity is the amount capable of reducing 2 μmol of cytochrome c min−1 at 25 °C. Purified his4-tagged recombinant wild-type LH (2 μM) was reduced VEGFR inhibitor with 50 mM freshly prepared

DTT for 1 h, treated with 200 mM iodomethane under a nitrogen-flushed atmosphere and left in the dark for a further 1 h. The unreduced enzyme was alkylated similarly as the control. The samples were passed through a Ni-agarose column (0.5 mL bed volume) to remove DTT from the treated sample. The control sample was eluted with 100 mM imidazole (pH 8) and 100 mM EDTA, but the Forskolin in vivo reduced/alkylated sample was eluted using 8 M urea and rapidly diluted with H2O to 0.8 M as it could not be eluted from the column under standard conditions. The activated LH was reduced with varying amounts of DTT (0–5 mM), and CdCl2 ranging from 0 to 25 mM was added to the preparations and incubated for 1 h. Excess DTT and CdCl2 were removed by dialysis of the protein solutions on 0.2-μm Millipore sterile filters in 20 mL 10 mM Tris–HCl (pH 8) for 1 h in a sterile Petri dish. Free thiol content estimation of lupanine hydroxylase in either native (wild type) or DTT-reduced state was published earlier in Stampolidis et al. (2009). Reaction of LH with

Ellman’s reagent occurred following the reduction of the thiol groups, but not in the unreduced state of the molecule, implying the potential presence of a disulphide bond. This initial observation formed the basis for the investigation presented in this paper. To determine whether the two Cys residues present in LH are disulphide bonded, a purified preparation of the recombinant wild-type enzyme was treated with iodomethane. Measurement Amylase of the specific activities of LH preparations of the reduced and unreduced alkylated enzyme had specific activities of 182 and 169 (± 5%) A555 min−1 mg−1 protein with 83% and 77% relative to control sample, respectively. However, the reduced and alkylated enzyme had a specific activity of 19 (± 5%) A555 min−1 mg−1 protein and only 9% activity relative to control sample (Table 1). The loss in activity of reduced/alkylated form indicated that Cys residues of LH must form a disulphide bond that could play a role in the activity and/or the stability of the enzyme.