Search PubMed⌕ Search

Biomedical subjects

J M Goodman

Publications and source records attributed to J M Goodman.

At least 73 records · Page 4Linked to original sources
PubMed1996

The targeting and assembly of peroxisomal proteins: some old rules do not apply.

Several proteins have been identified that catalyze the import of proteins into peroxisomes. Some recognize specific peroxisomal targeting sequences, but most probably work further downstream the import pathway. Recent evidence suggests that peroxisomal targeting and assembly do not follow the same rules as those for targeting and import into other organelles, such as the mitochondria and the endoplasmic reticulum, i.e. the import of unfolded proteins and subsequent folding within the organelle. Specifically, proteins may be translocated into the peroxisomal matrix in a folded or oligomerized state.

Biological Transport↗
PubMed1995

Cardiopulmonary function in adult patients late after Fontan repair.

OBJECTIVES: The clinical status and exercise assessment of adult patients late after the Fontan operation were reviewed to determine cardiovascular function. BACKGROUND: The Fontan operation is the final operation for many patients with tricuspid atresia or a single ventricle. Follow-up reports describe most patients to be in Canadian Cardiovascular Society functional class I or II. Objective measures of cardiac performance in the pediatric age group have shown significant dysfunction. METHODS: Forty-seven adult patients were seen late after the Fontan operation at the Toronto Congenital Cardiac Centre for Adults. Thirty of these underwent cycle ergometry to determine maximal exercise capacity. Maximal ventilation, maximal oxygen uptake and anaerobic threshold were determined from a ramp exercise protocol. Ejection fraction at rest and during exercise was measured with gated radionuclide angiography. Results were compared with those of eight normal volunteers. Results are given as mean +/- SD. RESULTS: Thirty patients underwent cardiopulmonary exercise testing 6.7 +/- 3.9 years after a first Fontan operation. Clinically 93% were in functional class I or II. The Fontan group patients had a significantly lower maximal work load (548 +/- 171 vs. 1,094 +/- 190 kilopond-meters, p < 0.00001), anaerobic threshold (11.2 +/- 2.9 vs. 23.6 +/- 4.6 ml/kg per min) and maximal oxygen consumption (14.8 +/- 4.5 vs. 42.1 +/- 10.0 ml/kg per min). Systemic ventricular ejection fraction was lower at rest (38 +/- 12% vs. 58 +/- 7%) and during exercise (40 +/- 15% vs. 70 +/- 8%). CONCLUSIONS: Despite a clinical impression of good function, by objective measures adult patients continue to have significant cardiovascular limitation late after the Fontan operation.

Adult↗
PubMed1995

Pmp27 promotes peroxisomal proliferation.

Peroxisomes perform many essential functions in eukaryotic cells. The weight of evidence indicates that these organelles divide by budding from preexisting peroxisomes. This process is not understood at the molecular level. Peroxisomal proliferation can be induced in Saccharomyces cerevisiae by oleate. This growth substrate is metabolized by peroxisomal enzymes. We have identified a protein, Pmp27, that promotes peroxisomal proliferation. This protein, previously termed Pmp24, was purified from peroxisomal membranes, and the corresponding gene, PMP27, was isolated and sequenced. Pmp27 shares sequence similarity with the Pmp30 family in Candida boidinii. Pmp27 is a hydrophobic peroxisomal membrane protein but it can be extracted by high pH, suggesting that it does not fully span the bilayer. Its expression is regulated by oleate. The function of Pmp27 was probed by observing the phenotype of strains in which the protein was eliminated by gene disruption or overproduced by expression from a multicopy plasmid. The strain containing the disruption (3B) was able to grow on all carbon sources tested, including oleate, although growth on oleate, glycerol, and acetate was slower than wild type. Strain 3B contained peroxisomes with all of the enzymes of beta-oxidation. However, in addition to the presence of a few modestly sized peroxisomes seen in a typical thin section of a cell growing on oleate-containing medium, cells of strain 3B also contained one or two very large peroxisomes. In contrast, cells in a strain in which Pmp27 was overexpressed contained an increased number of normal-sized peroxisomes. We suggest that Pmp27 promotes peroxisomal proliferation by participating in peroxisomal elongation or fission.

Acetates↗
PubMed1995

The Candida boidinii peroxisomal membrane protein Pmp30 has a role in peroxisomal proliferation and is functionally homologous to Pmp27 from Saccharomyces cerevisiae.

The mechanism of peroxisome proliferation is poorly understood. Candida boidinii is a methylotrophic yeast that undergoes rapid and massive peroxisome proliferation and serves as a good model system for this process. Pmp30A and Pmp30B (formerly designated Pmp31 and Pmp32, respectively) are two closely related proteins in a polyploid strain of this yeast that are strongly induced by diverse peroxisome proliferators such as methanol, oleate, and D-alanine. The function of these proteins is not understood. To study this issue, we used a recently described haploid strain (S2) of C. boidinii that can be manipulated genetically. We now report that strain S2 contains a single PMP30 gene very similar in sequence (greater than 93% identity at the DNA level) to PMP30A and PMP30B. When PMP30 was disrupted, cell growth on methanol was greatly inhibited, and cells grown in both methanol and oleate had fewer, larger, and more spherical peroxisomes than wild-type cells. A similar phenotype was recently described for Saccharomyces cerevisiae cultured on oleate in which PMP27, which encodes a protein of related sequence that is important for peroxisome proliferation, was disrupted. To determine whether Pmp27 is a functional homolog of Pmp30, gentle complementation was performed. PMP30A was expressed in the PMP27 disruptant of S. cerevisiae, and PMP27 was expressed in the PMP30 disruptant of C. boidinii S2. Complementation, in terms of both cell growth and organelle size, shape, and number, was successful in both directions, although reversion to a wild-type phenotype was only partial for the PMP30 disruptant. We conclude that these proteins are functional homologs and that both Pmp30 and Pmp27 have a direct role in proliferation and organelle size rather than a role in a specific peroxisomal metabolic pathway of substrate utilization.

Amino Acid Sequence↗
PubMed1995

Favorable left ventricular remodeling following large myocardial infarction by exercise training. Effect on ventricular morphology and gene expression.

Continued adverse remodeling of myocardium after infarction may lead to progressive ventricular dilation and heart failure. We tested the hypothesis that exercise training in a healed myocardial infarction-dysfunction rat model can favorably modify the adverse effects of ventricular remodeling including attenuation of abnormal myosin gene expression. Sprague-Dawley rats were subjected to either proximal LAD ligation or sham operation. At 5 wk after the operation, animals were randomly assigned to sedentary conditions or 6 wk of graduated swim training, creating four experimental groups: infarct sedentary (IS), infarct exercise (IE), sham sedentary (SS), and sham exercise (SE). At 11 wk all rats were sacrificed and analyzed. Compared to sedentary infarct controls, exercise training attenuated left ventricular (LV) dilation and allowed more hypertrophy of the non infarct wall. The exercise-trained hearts also showed a reduction in the estimated peak wall tension. Northern blot analysis showed an increase in beta-myosin heavy chain expression in the hearts of the sedentary infarction group soon after infarction when compared to sham controls. However, with exercise training, there was a significant attenuation of the beta-myosin heavy chain expression in the myocardium. Exercise training in a model of left ventricular dysfunction after healed myocardial infarction can improve the adverse remodeling process by attenuating ventricular dilation and reducing wall tension. The abnormal beta-myosin expression was also attenuated in the exercise trained group. This is evidence that abnormal gene expression following severe myocardial infarction dysfunction can be favorably modified by an intervention.

Animals↗
PubMed1994

The peroxisomal membrane proteins of Candida boidinii: gene isolation and expression.

Candida boidinii is a methylotrophic yeast in which several growth substrates can cause vigorous peroxisomal proliferation. While such diverse substrates as methanol, oleic acid and D-alanine induce different peroxisomal metabolic pathways, membranes seem to contain common abundant peroxisomal membrane proteins (PMPs). These proteins have been termed PMP31, PMP32 and PMP47. The gene encoding PMP47 has been previously cloned and analysed. We now report the isolation of a second PMP47 gene (or allele) as well as PMP31 and PMP32. PMP47A and PMP47B share 95% sequence identity at the amino acid level. PMP31 and PMP32 each contain 256 amino acids and are highly similar (97% identity) in protein sequence. Both PMP31 and PMP32 are predicted to span the membrane once or twice. All abundant PMPs of C. boidinii are basic in charge; they all have predicted isoelectric points above 10. RNAs corresponding to the PMP47s and to PMPs31-32 are strongly induced by methanol, oleic acid and D-alanine. While the PMP47s probably encode substrate carriers, the functions of PMP31 and PMP32 from C. boidinii are still unknown.

Alanine↗
PubMed1994

An internal region of the peroxisomal membrane protein PMP47 is essential for sorting to peroxisomes.

Targeting sequences on peroxisomal membrane proteins have not yet been identified. We have attempted to find such a sequence within PMP47, a protein of the methylotrophic yeast, Candida boidinii. This protein of 423 amino acids shows sequence similarity with proteins in the family of mitochondrial carrier proteins. As such, it is predicted to have six membrane-spanning domains. Protease susceptibility experiments are consistent with a six-membrane-spanning model for PMP47, although the topology for the peroxisomal protein is inverted compared with the mitochondrial carrier proteins. PMP47 contains two potential peroxisomal targeting sequences (PTS1), an internal SKL (residues 320-322) and a carboxy terminal AKE (residues 421-423). Using a heterologous in vivo sorting system, we show that efficient sorting occurs in the absence of both sequences. Analysis of PMP47-dihydrofolate reductase (DHFR) fusion proteins revealed that amino acids 1-199 of PMP47, which contain the first three putative membrane spans, do not contain the necessary targeting information, whereas a fusion with amino acids 1-267, which contains five spans, is fully competent for sorting to peroxisomes. Similarly, a DHFR fusion construct containing residues 268-423 did not target to peroxisomes while residues 203-420 appeared to sort to that organelle, albeit at lower efficiency than the 1-267 construct. However, DHFR constructs containing only amino acids 185-267 or 203-267 of PMP47 were not found to be associated with peroxisomes. We conclude that amino acids 199-267 are necessary for peroxisomal targeting, although additional sequences may be required for efficient sorting to, or retention by, the organelles.

Amino Acid Sequence↗
PubMed1994

An oligomeric protein is imported into peroxisomes in vivo.

The mechanism of translocation of peroxisomal proteins from the cytoplasm into the matrix is largely unknown. We have been studying this problem in yeast. We show that the peroxisomal targeting sequences SKL or AKL, with or without a spacer of nine glycines (G9), are sufficient to target chloramphenicol acetyltransferase (CAT) to peroxisomes of Saccharomyces cerevisiae in vivo. The mature form of CAT is a homotrimer, and complete trimerization of CAT was found to occur within a few minutes of synthesis. In contrast, import, measured by immunoelectron microscopy and organellar fractionation, occurred over several hours. To confirm that import of preassembled CAT trimers was occurring, we co-expressed CAT-G9-AKL with CAT lacking a peroxisomal targeting sequence but containing a hemagglutinin-derived epitope tag (HA-CAT). We found that HA-CAT was not imported unless it was co-expressed with CAT-G9-AKL. Both proteins were released from the organelles under mild conditions (pH 8.5) that released other matrix proteins, indicating that import had occurred. These results strongly suggested that HA-CAT was imported as a heterotrimer with CAT-G9-AKL. The process of oligomeric import also occurs in animal cells. When HA-CAT was co-expressed with CAT-G9-AKL in CV-1 cells, HA-CAT co-localized with peroxisomes but was cytoplasmic when expressed alone. It is not clear whether the import of globular proteins into peroxisomes occurs through peroxisomal membrane pores or involves membrane internalization. Both possibilities are discussed.

Amino Acid Sequence↗
PubMed1994

Measurement of left ventricular function during arm ergometry using the VEST nuclear probe.

A chest-mounted left ventricular (LV) nuclear probe (VEST) for use during arm and leg ergometry is presented, with a discussion of the validity and reproducibility of LV function measures at rest and exercise. During both arm and leg ergometry in trained subjects, transient changes in LV function/volumes were observed. LV ejection fraction and relative end-systolic and end-diastolic volumes were 25 to 30% less with the arms versus the legs, agreeing with data from other studies using conventional techniques. At peak exercise with both limbs, LV ejection fraction and relative LV end-systolic volume increased, followed by immediate postexercise normalization. The effect was greatest with the arms and reflects the effect of high intramuscular and arterial pressures generated during arm cranking, leading to increased LV afterloading. The VEST permits rapid and noninvasive assessment of LV function during arm exercise, avoiding the limitations of other techniques.

Blood Pressure↗
PubMed1993

Expression and targeting of a 47 kDa integral peroxisomal membrane protein of Candida boidinii in wild type and a peroxisome-deficient mutant of Hansenula polymorpha.

A 47 kDa integral peroxisomal membrane protein (PMP47) of Candida boidinii was expressed in wild type (WT) and a temperature-sensitive (Ts6) peroxisome-deficient (per) mutant of Hansenula polymorpha. The subcellular location of PMP47 appeared to be dependent on the level of expression. At low expression levels PMP47 was sorted to the peroxisomal membrane; however, in Ts6 cells grown at restrictive temperatures (which lack intact peroxisomes) PMP47 was solely located in small cytosolic aggregates, together with homologous H. polymorpha PMP's. At enhanced expression levels, however, part of the protein also became incorporated into mitochondria, both in transformed WT and Ts6 cells.

Blotting, Western↗
PubMed1993

Clinical assessment of anaerobic isolates from blood cultures.

Patients at two tertiary-care medical centers were evaluated to determine the clinical significance of anaerobic isolates from their blood specimens and to identify whether aerobic and/or anaerobic conditions were necessary for the detection of Streptococcus pneumoniae isolates. Significant anaerobes were isolated from only 0.1% and 0.4% of all blood cultures collected. The majority of patients with significant anaerobes had clinical conditions in which anaerobes are known to cause infections. Of the S. pneumoniae organisms, 83% were isolated only from the aerobic bottles of a blood culture set. These data lend support to the recommendations for the selective ordering of anaerobic blood cultures without compromising the isolation of S. pneumoniae.

Adolescent↗
PubMed1993

Atrial natriuretic peptide during acute and prolonged exercise in well-trained men.

In this study, cardiac performance and circulating immunoreactive atrial natriuretic peptide (Ir-ANP) were measured during sustained exercise to examine the relationship between cardiac filling, heart rate and circulating ANP. Fifteen well-trained male subjects (mean age = 27.6 +/- 1 years) were studied during two exercise sessions. Initially, graded exercise to maximum was performed to determine maximal oxygen consumption (VO2max). On a separate occasion subjects performed 150 minutes of continuous exercise at 70-74% of the maximal heart rate, with radionuclide angiography performed at rest and every 50 minutes, in conjunction with Ir-ANP, blood pressure and heart rate measurements. During maximal exercise subjects reached a VO2max of 47.9 +/- 2.1 ml/kg/min. Ir-ANP increased from 9.8 +/- 1.1 pg/ml at rest to 45.2 +/- 6.6 pg/ml at maximal exercise. Ir-ANP remained elevated (28.9 +/- 3.4 pg/ml) 3 minutes after exercise (p < 0.05). During prolonged exercise, left ventricular end-diastolic filling decreased significantly between 50 and 150 minutes (p < 0.05), with progressive rise in heart rate observed between 50 (141 +/- 8 beats/minute) and 150 minutes (154 +/- 3 beats/min; p < 0.05). No change in left ventricular end-systolic counts, or arterial blood pressure were observed throughout the exercise period. Ir-ANP was elevated significantly during prolonged exercise, increasing from 9.4 +/- 1.7 at rest, to 19.2 +/- 3.4 after 50 minutes, 17.8 +/- 4.3 pg/ml after 100 minutes of exercise (p < 0.05), and 23.5 +/- 2.1 pg/ml by 150 minutes of exercise. A weak correlation (r = 0.33) was observed between heart rate and Ir-ANP throughout the exercise session.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

Specific cross-linking of the proline isomerase cyclophilin to a non-proline-containing peptide.

A peptide corresponding to an efficient peroxisomal targeting sequence, the carboxy terminal 12 amino acids of PMP20 from Candida boidinii, was employed as an affinity ligand to search for a peroxisomal targeting receptor. Two proteins from yeast extracts with apparent molecular masses of 20 and 80 kDa were detected by chemical cross-linking to radioiodinated peptide. Both proteins were present in cytosolic supernatants. The 20-kDa species did not cross-link to a control peptide with reversed sequence, whereas the 80-kDa protein cross-linked to both peptides. The cross-linking assay was used to purify the 20-kDa protein from Saccharomyces cerevisiae. Partial protein sequencing identified this protein as cyclophilin, the product of the CYP1 gene. This protein, a peptidyl-prolyl cis-trans isomerase, is the yeast homologue of the protein that mediates the immunosuppressant effects of the drug cyclosporin A (CsA). Cross-linking of peptide to cyclophilin was inhibited by CsA. The cross-linking of cyclophilin to the PMP20-derived peptide was unanticipated because the peptide contains no prolines. The CYP1-encoded protein was not required to target proteins to peroxisomes because this organelle appeared to be assembled normally in a CYP1-disrupted strain. Furthermore, the final three amino acids of the peptide, which are critical for peroxisomal sorting, were not required for cross-linking to cyclophilin. We conclude that either cyclophilin is playing a nonessential facilitating role in peroxisomal targeting or that the interaction of the targeting peptide to cyclophilin is mimicking an interaction with an unidentified substrate or effector of cyclophilin.

Amino Acid Isomerases↗
PubMed1993

Vascular conductance and aerobic power in sedentary and active subjects and heart failure patients.

The relationship between peak aerobic power and a strain-gauge determination of local skeletal muscle vascular conductance 10-13 s after calf exercise to fatigue was examined in 21 middle-aged adults (age 38.1 +/- 2.5 yr): seven physically active men (A), seven sedentary men (S), and six men and one woman with compensated idiopathic heart failure (HF). The three subgroups were chosen as differing widely in peak O2 intake [48.2 +/- 1.2, 32.9 +/- 1.6, and 16.1 +/- 1.3 (SE) ml.kg-1 x min-1, respectively]. Calf vascular conductance showed a gradation with aerobic power: 64.8 +/- 3.8, 40.7 +/- 4.3, and 30.7 +/- 6.1 (SE) ml/min local flow per 10 liters of tissue per unit of blood pressure. There was a strong positive correlation between O2 intake and vascular conductance for the overall group (VO2 = 0.614 G + 3.5; r = 0.75, P < 0.001) and for the 14 normal subjects (VO2 = 0.377 G + 20.6; r = 0.74, P < 0.002). The mean conductance was smaller in HF (P < 0.001), with no significant slope in relation to O2 intake. There was no relationship between the resting cardiac ejection fraction [74.4 +/- 4.1% (SE) for A, 74.3 +/- 4.2% for S, and 25.8 +/- 5.2% for HF] and either peak aerobic power or calf vascular conductance. We conclude that peak aerobic power is strongly associated with local vascular conductance during peripherally limited exercise involving the calf muscles of one leg and that vascular conductance is particularly low in subjects with compensated idiopathic heart failure.

Adult↗
PubMed1992

Ligand atom partial charges assignment for complementary electrostatic potentials.

The design of molecules to fit into the active site of receptors is a rapidly developing area of pharmacology and medicinal chemistry. A good ligand needs a suitable geometry and also appropriate electrostatic properties. The electrostatic properties of the ligand should complement those of the receptor. We present a method for the assignment of atom-centred point charges for a ligand, based on the electrostatic potential of the receptor. These point charges are chosen to give the best possible complementarity to the receptor electrostatic potential over the van der Waals surface of the ligand. We demonstrate that point charges can be chosen to give good electrostatic complementarity, and suggest that a molecule with similar electrostatic properties should bind well to the receptor.

Binding Sites↗
PubMed1992

Development of multipurpose peroxisomes in Candida boidinii grown in oleic acid-methanol limited continuous cultures.

We have studied the development and metabolic significance of peroxisomes in the yeast Candida boidinii following adaptation of the organism to cultivation conditions which require the simultaneous presence and activity of two independent peroxisome-mediated pathways for growth. After the addition of methanol to oleic acid-grown cells at late exponentional growth, a number of new small peroxisomes developed which, apart from the presence of beta-oxidation enzymes, were characterized by the presence of enzymes involved in methanol metabolism (alcohol oxidase and dihydroxyacetone synthase). The latter proteins, however, were absent in the larger organelles which were originally present in the oleic acid-grown cells prior to the addition of methanol and which contained only enzymes of the beta-oxidation pathway. Subsequent experiments on cells from continuous cultures grown on a mixture of oleic acid and methanol at steady-state conditions revealed that both the enzymes of the beta-oxidation pathway and those involved in methanol metabolism were found in one and the same compartment. Thus, under these conditions the cells contained peroxisomes which were concurrently involved in the metabolism of two different carbon sources simultaneously used for growth. Our results indicated that the heterogeneity in the peroxisomal population of a single cell, observed in the transient state following the addition of methanol, is only temporary and due to heterogeneity among these organelles with respect to their capacity to incorporate newly synthesized matrix proteins.

Acetyl-CoA C-Acetyltransferase↗