22 research outputs found
Molecular Evolutionary Analysis of the Thiamine-Diphosphate-Dependent Enzyme, Transketolase
Members of the transketolase group of thiamine-
diphosphate-dependent enzymes from 17 different
organisms including mammals, yeast, bacteria, and
plants have been used for phylogenetic reconstruction.
Alignment of the amino acid and DNA sequences for 21
transketolase enzymes and one putative transketolase reveals
a number of highly conserved regions and invariant
residues that are of predicted importance for enzyme
activity, based on the crystal structure of yeast transketolase.
One particular sequence of 36 residues has some
similarities to the nucleotide-binding motif and we designate
it as the transketolase motif. We report further
evidence that the recP protein from Streptococcus pneumoniae
might be a transketolase and we list a number of
invariant residues which might be involved in substrate
binding. Phylogenies derived from the nucleotide and the
amino acid sequences by various methods show a conventional
clustering for mammalian, plant, and gramnegative
bacterial transketolases. The branching order of
the gram-positive bacteria could not be inferred reliably.
The formaldehyde transketolase (sometimes known as
dihydroxyacetone synthase) of the yeast Hansenula polymorpha
appears to be orthologous to the mammalian
enzymes but paralogous to the other yeast transketolases.
The occurrence of more than one transketolase gene in
some organisms is consistent with several gene duplications.
The high degree of similarity in functionally important
residues and the fact that the same kinetic mechanism
is applicable to all characterized transketolase
enzymes is consistent with the proposition that they are
all derived from one common ancestral gene. Transketolase
appears to be an ancient enzyme that has evolved
slowly and might serve as a model for a molecular clock,
at least within the mammalian clade
Molecular Evolutionary Analysis of the Thiamine-Diphosphate-Dependent Enzyme, Transketolase
Members of the transketolase group of thiamine-
diphosphate-dependent enzymes from 17 different
organisms including mammals, yeast, bacteria, and
plants have been used for phylogenetic reconstruction.
Alignment of the amino acid and DNA sequences for 21
transketolase enzymes and one putative transketolase reveals
a number of highly conserved regions and invariant
residues that are of predicted importance for enzyme
activity, based on the crystal structure of yeast transketolase.
One particular sequence of 36 residues has some
similarities to the nucleotide-binding motif and we designate
it as the transketolase motif. We report further
evidence that the recP protein from Streptococcus pneumoniae
might be a transketolase and we list a number of
invariant residues which might be involved in substrate
binding. Phylogenies derived from the nucleotide and the
amino acid sequences by various methods show a conventional
clustering for mammalian, plant, and gramnegative
bacterial transketolases. The branching order of
the gram-positive bacteria could not be inferred reliably.
The formaldehyde transketolase (sometimes known as
dihydroxyacetone synthase) of the yeast Hansenula polymorpha
appears to be orthologous to the mammalian
enzymes but paralogous to the other yeast transketolases.
The occurrence of more than one transketolase gene in
some organisms is consistent with several gene duplications.
The high degree of similarity in functionally important
residues and the fact that the same kinetic mechanism
is applicable to all characterized transketolase
enzymes is consistent with the proposition that they are
all derived from one common ancestral gene. Transketolase
appears to be an ancient enzyme that has evolved
slowly and might serve as a model for a molecular clock,
at least within the mammalian clade
Fostering Equity and Diversity in the Nova Scotia Legal Profession
The Province of Nova Scotia has, for many years, attempted, through a variety of means, to address issues of diversity and affirmative action. However, despite the lessons of history there are still those who question the need for programs and policies that promote, encourage and enforce equality. Even though significant advances have been made on many fronts Nova Scotia continues to struggle with issues of inequality. As with many problems faced by society acknowledging the existence of the problem is the first step towards developing solutions
Mortality from gastrointestinal congenital anomalies at 264 hospitals in 74 low-income, middle-income, and high-income countries: a multicentre, international, prospective cohort study
Summary
Background Congenital anomalies are the fifth leading cause of mortality in children younger than 5 years globally.
Many gastrointestinal congenital anomalies are fatal without timely access to neonatal surgical care, but few studies
have been done on these conditions in low-income and middle-income countries (LMICs). We compared outcomes of
the seven most common gastrointestinal congenital anomalies in low-income, middle-income, and high-income
countries globally, and identified factors associated with mortality.
Methods We did a multicentre, international prospective cohort study of patients younger than 16 years, presenting to
hospital for the first time with oesophageal atresia, congenital diaphragmatic hernia, intestinal atresia, gastroschisis,
exomphalos, anorectal malformation, and Hirschsprung’s disease. Recruitment was of consecutive patients for a
minimum of 1 month between October, 2018, and April, 2019. We collected data on patient demographics, clinical
status, interventions, and outcomes using the REDCap platform. Patients were followed up for 30 days after primary
intervention, or 30 days after admission if they did not receive an intervention. The primary outcome was all-cause,
in-hospital mortality for all conditions combined and each condition individually, stratified by country income status.
We did a complete case analysis.
Findings We included 3849 patients with 3975 study conditions (560 with oesophageal atresia, 448 with congenital
diaphragmatic hernia, 681 with intestinal atresia, 453 with gastroschisis, 325 with exomphalos, 991 with anorectal
malformation, and 517 with Hirschsprung’s disease) from 264 hospitals (89 in high-income countries, 166 in middleincome
countries, and nine in low-income countries) in 74 countries. Of the 3849 patients, 2231 (58·0%) were male.
Median gestational age at birth was 38 weeks (IQR 36–39) and median bodyweight at presentation was 2·8 kg (2·3–3·3).
Mortality among all patients was 37 (39·8%) of 93 in low-income countries, 583 (20·4%) of 2860 in middle-income
countries, and 50 (5·6%) of 896 in high-income countries (p<0·0001 between all country income groups).
Gastroschisis had the greatest difference in mortality between country income strata (nine [90·0%] of ten in lowincome
countries, 97 [31·9%] of 304 in middle-income countries, and two [1·4%] of 139 in high-income countries;
p≤0·0001 between all country income groups). Factors significantly associated with higher mortality for all patients
combined included country income status (low-income vs high-income countries, risk ratio 2·78 [95% CI 1·88–4·11],
p<0·0001; middle-income vs high-income countries, 2·11 [1·59–2·79], p<0·0001), sepsis at presentation (1·20
[1·04–1·40], p=0·016), higher American Society of Anesthesiologists (ASA) score at primary intervention
(ASA 4–5 vs ASA 1–2, 1·82 [1·40–2·35], p<0·0001; ASA 3 vs ASA 1–2, 1·58, [1·30–1·92], p<0·0001]), surgical safety
checklist not used (1·39 [1·02–1·90], p=0·035), and ventilation or parenteral nutrition unavailable when needed
(ventilation 1·96, [1·41–2·71], p=0·0001; parenteral nutrition 1·35, [1·05–1·74], p=0·018). Administration of
parenteral nutrition (0·61, [0·47–0·79], p=0·0002) and use of a peripherally inserted central catheter (0·65
[0·50–0·86], p=0·0024) or percutaneous central line (0·69 [0·48–1·00], p=0·049) were associated with lower mortality.
Interpretation Unacceptable differences in mortality exist for gastrointestinal congenital anomalies between lowincome,
middle-income, and high-income countries. Improving access to quality neonatal surgical care in LMICs will
be vital to achieve Sustainable Development Goal 3.2 of ending preventable deaths in neonates and children younger
than 5 years by 2030
number I, 95-98 FEBS 10579 Q 1992 Federation oF European Uiochcmical Sociciics
A trypiophan residue at position 487 in Z.wromomzs mohilis pyruvatc drcarboxylase was altered to leucine by sitcdirccted mutagenesis. This modified Z. rtrobili.~ pyruvatc decarboxylase was active when expressed in Escfwichiu c&and had unchanged kinaics towards pyruvatc. The em/me showed a decreasti allinity For the cohclors with the half-saturating concentrations increasing from 0.64 to 9.0jtM for thiamin diphosphatc and from 4.21 to 45 PM for My". Unlike the wild-type enzyme, there was little quenching of tryptophan fluomccnse upon adding cofactors to this modified form. The data suggest that tryptophan-487 is close to the cofactor binding site but is not required absolutely for pyruvate dccarboxylase activity. SubstituGon of asparagine, threoninc or glycine for aspartate-440, a r&due which is conserved between many thinmin diphosphatedependent enzymes, completely abolishes enzyme activity. Pyruvute decarboxylase; Site-directed mutagenesis: Thiamin diphosphate binding; ZJVHONW~I~ rtrubili
The role of residues glutamate-50 and phenylalanine-496 in Zymomonas mobilis pyruvate decarboxylase
Molecular Evolutionary Analysis of the Thiamine-Diphosphate-Dependent Enzyme, Transketolase
Members of the transketolase group of thiamine-
diphosphate-dependent enzymes from 17 different
organisms including mammals, yeast, bacteria, and
plants have been used for phylogenetic reconstruction.
Alignment of the amino acid and DNA sequences for 21
transketolase enzymes and one putative transketolase reveals
a number of highly conserved regions and invariant
residues that are of predicted importance for enzyme
activity, based on the crystal structure of yeast transketolase.
One particular sequence of 36 residues has some
similarities to the nucleotide-binding motif and we designate
it as the transketolase motif. We report further
evidence that the recP protein from Streptococcus pneumoniae
might be a transketolase and we list a number of
invariant residues which might be involved in substrate
binding. Phylogenies derived from the nucleotide and the
amino acid sequences by various methods show a conventional
clustering for mammalian, plant, and gramnegative
bacterial transketolases. The branching order of
the gram-positive bacteria could not be inferred reliably.
The formaldehyde transketolase (sometimes known as
dihydroxyacetone synthase) of the yeast Hansenula polymorpha
appears to be orthologous to the mammalian
enzymes but paralogous to the other yeast transketolases.
The occurrence of more than one transketolase gene in
some organisms is consistent with several gene duplications.
The high degree of similarity in functionally important
residues and the fact that the same kinetic mechanism
is applicable to all characterized transketolase
enzymes is consistent with the proposition that they are
all derived from one common ancestral gene. Transketolase
appears to be an ancient enzyme that has evolved
slowly and might serve as a model for a molecular clock,
at least within the mammalian clade
Molecular Evolutionary Analysis of the Thiamine-Diphosphate-Dependent Enzyme, Transketolase
Members of the transketolase group of thiamine-
diphosphate-dependent enzymes from 17 different
organisms including mammals, yeast, bacteria, and
plants have been used for phylogenetic reconstruction.
Alignment of the amino acid and DNA sequences for 21
transketolase enzymes and one putative transketolase reveals
a number of highly conserved regions and invariant
residues that are of predicted importance for enzyme
activity, based on the crystal structure of yeast transketolase.
One particular sequence of 36 residues has some
similarities to the nucleotide-binding motif and we designate
it as the transketolase motif. We report further
evidence that the recP protein from Streptococcus pneumoniae
might be a transketolase and we list a number of
invariant residues which might be involved in substrate
binding. Phylogenies derived from the nucleotide and the
amino acid sequences by various methods show a conventional
clustering for mammalian, plant, and gramnegative
bacterial transketolases. The branching order of
the gram-positive bacteria could not be inferred reliably.
The formaldehyde transketolase (sometimes known as
dihydroxyacetone synthase) of the yeast Hansenula polymorpha
appears to be orthologous to the mammalian
enzymes but paralogous to the other yeast transketolases.
The occurrence of more than one transketolase gene in
some organisms is consistent with several gene duplications.
The high degree of similarity in functionally important
residues and the fact that the same kinetic mechanism
is applicable to all characterized transketolase
enzymes is consistent with the proposition that they are
all derived from one common ancestral gene. Transketolase
appears to be an ancient enzyme that has evolved
slowly and might serve as a model for a molecular clock,
at least within the mammalian clade
Establishment of a Functional Genomics Platform for Leifsonia xyli subsp. xyli
Leifsonia xyli subsp. xyli, the causal agent of ratoon stunting disease in sugarcane, is a xylem-limited, nutritionally fastidious, slow growing, gram-positive coryneform bacterium. Because of the difficulties in growing this bacterium in pure culture, little is known about the molecular mechanisms of pathogenesis. Currently, the genome sequence of L. xyli subsp. xyli is being completed by the Agronomical and Environmental Genomes group from the Organization for Nucleotide Sequencing and Analysis in Brazil. To complement this work, we produced 712 LxxTn4431 transposon mutants and sequenced flanking regions from 383 of these, using a rapid polymerase chain reaction-based approach. Tn4431 insertions appeared to be widespread throughout the L. xyli subsp. xyli genome; however, there were regions that had significantly higher concentrations of insertions. The Tn4431 mutant library was screened for individuals unable to colonize sugarcane, and one noncolonizing mutant was found. The mutant contained a transposon insertion disrupting two open reading frames (ORF), one of which had homology to an integral membrane protein from Mycobacterium leprae. Sequencing of the surrounding regions revealed two operons, pro and cyd, both of which are believed to play roles in disease. Complementation studies were carried out using the noncolonizing LxxTn4431 mutant. The noncolonizing mutant was transformed with a cosmid containing 40 kbp of wild-type sequence, which included the two ORF disrupted in the mutant, and several transformants were subsequently able to colonize sugarcane. However, analysis of each of these transformants, before and after colonization, suggests that they have all undergone various recombinant events, obscuring the roles of these ORF in L. xyli subsp. xyli pathogenesis