Identification, cloning, and characterization of a Lactococcus lactis branched-chain alpha-keto acid decarboxylase involved in flavor formation

The biochemical pathway for formation of branched-chain aldehydes, which are important flavor compounds derived from proteins in fermented dairy products, consists of a protease, peptidases, a transaminase, and a branched-chain alpha-keto acid decarboxylase (KdcA). The activity of the latter enzyme has been found only in a limited number of Lactococcus lactis strains. By using a random mutagenesis approach, the gene encoding KdcA in L. lactis B1157 was identified. The gene for this enzyme is highly homologous to the gene annotated ipd, which encodes a putative indole pyruvate decarboxylase, in L. lactis IL1403. Strain IL1403 does not produce KdcA, which could be explained by a 270-nucleotide deletion at the 3' terminus of the ipd gene encoding a truncated nonfunctional decarboxylase. The kdcA gene was overexpressed in L. lactis for further characterization of the decarboxylase enzyme. Of all of the potential substrates tested, the highest activity was observed with branched-chain alpha-keto acids. Moreover, the enzyme activity was hardly affected by high salinity, and optimal activity was found at pH 6.3, indicating that the enzyme might be active under cheese ripening conditions.     

Gene expression and digit homology in the chicken embryo wing

The bird wing is of special interest to students of homology and avian evolution. Fossil and developmental data give conflicting indications of digit homology if a pentadactyl "archetype" is assumed. Morphological signs of a vestigial digit I are seen in bird embryos, but no digit-like structure develops in wild-type embryos. To examine the developmental mechanisms of digit loss, we studied the expression of the high-mobility group box containing Sox9 gene, and bone morphogenetic protein receptor 1b (bmpR-1b)-markers for precondensation and prechondrogenic cells, respectively. We find an elongated domain of Sox9 expression, but no bmpR-1b expression, anterior to digit II. We interpret this as a digit I domain that reaches precondensation, but not condensation or precartilage stages. It develops late, when the tissue in which it is lodged is being remodeled. We consider these findings in the light of previous Hoxd-11 misexpression studies. Together, they suggest that there is a digit I vestige in the wing that can be rescued and undergo development if posterior patterning cues are enhanced. We observed Sox9 expression in the elusive "element X" that is sometimes stated to represent a sixth digit. Indeed, incongruity between digit domains and identities in theropods disappears if birds and other archosaurs are considered primitively polydactyl. Our study provides the first gene expression evidence for at least five digital domains in the chick wing. The failure of the first to develop may be plausibly linked to attenuation of posterior signals.     

Can folic acid protect against congenital heart defects in down syndrome?

BACKGROUND: Several studies have suggested a protective effect of folic acid (FA) on congenital heart anomalies. Down syndrome (DS) infants are known to have a high frequency of heart anomalies. Not all children with DS suffer from heart anomalies, which raises the question whether maternal factors might affect the risk of these anomalies. Our objectives were to investigate whether first-trimester FA use protects against heart anomalies among DS children. METHODS: Women with liveborn DS children participating in the Slone Epidemiology Center Birth Defects Study between 1976 and 1997 were included. We performed case-control analyses using DS, with heart anomalies as cases and DS, without heart anomalies as controls. Subanalyses were performed for defects that have been associated with FA in non-DS populations (conotruncal, ventricular septal [VSD]) and for those that are associated with DS (ostium secundum type atrial septal defects [ASD] and endocardial cushion defects [ECD]). Exposure was defined as the use of any FA-containing product for an average of at least 4 days per week during the first 12 weeks of pregnancy, whereas no exposure was defined as no use of FA in these 12 weeks. RESULTS: Of the 223 cases, 110 (49%) were exposed versus 84 (46%) of the 184 controls. After adjustment for possible confounders, no protective effect of FA was found on heart anomalies overall (OR 0.95, 95% CI: 0.61-1.47) nor separately for conotruncal defects, VSDs, ASDs, or ECDs. CONCLUSIONS: Our study does not show a protective effect of FA on heart anomalies among infants with DS.     

Observation of two modes of inhibition of human microsomal prostaglandin E synthase 1 by the cyclopentenone 15-deoxy-Δ(12,14)-prostaglandin J(2)

Microsomal prostaglandin E synthase 1 (MPGES1) is an enzyme that produces the pro-inflammatory molecule prostaglandin E(2) (PGE(2)). Effective inhibitors of MPGES1 are of considerable pharmacological interest for the selective control of pain, fever, and inflammation. The isoprostane, 15-deoxy-Δ(12,14)-prostaglandin J(2) (15d-PGJ(2)), a naturally occurring degradation product of prostaglandin D(2), is known to have anti-inflammatory properties. In this paper, we demonstrate that 15d-PGJ(2) can inhibit MPGES1 by covalent modification of residue C59 and by noncovalent inhibition through binding at the substrate (PGH(2)) binding site. The mechanism of inhibition is dissected by analysis of the native enzyme and the MPGES1 C59A mutant in the presence of glutathione (GSH) and glutathione sulfonate. The location of inhibitor adduction and noncovalent binding was determined by triple mass spectrometry sequencing and with backbone amide H/D exchange mass spectrometry. The kinetics, regiochemistry, and stereochemistry of the spontaneous reaction of GSH with 15d-PGJ(2) were determined. The question of whether the anti-inflammatory properties of 15d-PGJ(2) are due to inhibition of MPGES1 is discussed.     

Prevalence of genital HPV infections and HPV serology in adolescent girls,prior to vaccination

Introduction: Monitoring the prevalence of type-specific HPV-DNA infections before and shortly after introduction of routine HPV vaccination offers the opportunity to evaluate early effects of the vaccination program. With this aim a cohort study was set up of 14- to 16-year-old girls eligible for HPV vaccination in the Netherlands. Annually, HPV-DNA and antibody status in vaginal self-samples and in serum respectively, will be studied among vaccinated (58%) and unvaccinated girls (42%). Here we present baseline data on vaginal HPV-DNA status in relation to serum antibodies. Methods: The 1800 enrolled girls filled out an internet-based questionnaire and provided a vaginal self-sample for genotype specific HPV-DNA detection using SPF₁₀ PCR amplification and reverse line probe hybridization. Furthermore, 64% of the girls provided a blood sample for HPV antibody analysis. IgG antibodies against virus-like particles were determined for 7 HPV genotypes. Results: At baseline, type-specific HPV-DNA was detected in 4.4% (n = 79) of the 1800 girls: 2.7% (n = 49) concerned a high risk HPV type (hrHPV-DNA). The three most common types were HPV type 16, 18 and 51 (40%). Out of the hrHPV-DNA positive girls, 32% was seropositive vs. 12% in HPV-DNA negative girls (p < 0.001). Risk factors independently associated with hrHPV-DNA infection among the sexually active girls were age >15 years vs. 14–15 years (OR = 2.6 (1.2–5.9)), age of sexual debut <14 vs. above 14 years (OR = 3.0 (1.1–8.2)), total number of lifetime partners above two vs. less than two partners (OR = 3.2 (1.3–8.0)) and age of partner >17 vs. under 17 years (OR = 4.2 (1.5–13.0)). Conclusion: A low hrHPV-DNA prevalence was found in the adolescent girls. The observed vs. expected age-related increase in HPV-DNA prevalence in this cohort in the coming years (with increased sexual activity) will provide understanding of the effect of HPV vaccination. Furthermore, this cohort study will offer the opportunity to improve knowledge of antibody responses following natural infection and vaccination.