Crystallization and structure determination of bovine profilin at 2.0 A resolution

Profilin regulates the behavior of the eukaryotic microfilament system through its interaction with non-filamentous actin. It also binds several ligands, including poly(L-proline) and the membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). Bovine profilin crystals (space group C2; a = 69.15 A, b = 34.59 A, c = 52.49 A; alpha = gamma = 90 degrees, beta = 92.56 degrees) were grown from a mixture of poly(ethylene glycol) 400 and ammonium sulfate. X-ray diffraction data were collected on an imaging plate scanner at the DORIS storage ring (DESY, Hamburg), and were phased by molecular replacement, using a search model derived from the 2.55 A structure of profilin complexed to beta-actin. The refined model of bovine profilin has a crystallographic R-factor of 16.5% in the resolution range 6.0 to 2.0 A and includes 128 water molecules, several of which form hydrogen bonds to stabilize unconventional turns. The structure of free bovine profilin is similar to that of bovine profilin complexed to beta-actin, and C alpha atoms from the two structures superimpose with an r.m.s. deviation of 1.25 A. This value is reduced to 0.51 A by omitting Ala1 and the N-terminal acetyl group, which lie at a profilin-actin interface in crystals of the complex. These residues display a strained conformation in crystalline profilin-actin but may allow the formation of a hydrogen bond between the N-acetyl carbonyl group of profilin and the phenol hydroxyl group of Tyr188 in actin. Several other actin-binding residues of profilin show different side-chain rotomer conformations in the two structures. The polypeptide fold of bovine profilin is generally similar to those observed by NMR for profilin from other sources, although the N terminus of Acanthamoeba profilin isoform I lies in a distorted helix and the C-terminal helix is less tilted with respect to the strands in the central beta-pleated sheet than is observed in bovine profilin. The majority of the aromatic residues in profilin are exposed to solvent and lie in either of two hydrophobic patches, neither of which takes part in an interface with actin. One of these patches is required for binding poly(L-proline) and contains an aromatic cluster comprising the highly conserved residues Trp3, Tyr6, Trp31 and Tyr139. In forming this cluster, Trp31 adopts a sterically strained rotamer conformation.(ABSTRACT TRUNCATED AT 400 WORDS)     

Homozygous protein C deficiency can be detected by prenatal diagnosis

Homozygous protein C deficiency (HPCD) with purpura fulminans is a rare condition with an estimated incidence of 1-2 per 400,000 births. About 20 case reports have appeared since the first one was published in 1983. HPCD provides an excellent illustration of the fundamental importance of the protein C anticoagulant pathway. This severe coagulopathy results in serious organ damage, often already in utero, and without treatment it is incompatible with life. The treatment options include fresh frozen plasma and protein C concentrate in the acute phase, followed by oral anticoagulant therapy. Over 160 different point mutations in the protein C gene have been identified in recent years, offering new possibilities for prenatal diagnosis. The article describes the case of a family who lost two children with congenital HPCD. But where the specific point mutation was identified thus enabling prenatal diagnosis to be performed in a subsequent pregnancy.     

Significance of IgG for the activity of factor XII measured in human plasma

The plasma level of factor XII (FXII) was measured in samples from healthy young men. The activated contact factor was assayed as prekallikrein activator (PKA), as S-2222 amidase, and in radial immunodiffusion tests. By removing the bulk of IgG on protein G columns before the activation procedure, the functional activities increased to about 135%. In such test preparations, PAGE immunoblot experiments with polyclonal antibodies against FXII showed, in addition to FXIIa (80 kD), a double band with a molecular weight of about 46 kD. This protein could also be detected with a light-chain-specific monoclonal antibody to FXII, but not with such an antibody directed against its heavy chain. The 46-kD band was also observed in plasma deficient in FXII. The amidase assays indicated that the minor part of FXIIa was present in some kind of association with another protease. To obtain a correct estimation of total FXIIa in the amidase assays a sufficiently high level of FXI was required compared to that of FXII. The PKA assays were generally carried out with a prekallikrein (PK) substrate containing IgG. By replacing this substrate by PK free from IgG additional PKA activity was observed, the activity appearing also in plasma deficient in FXII.     

Effectiveness of a worksite smoking cessation program in the military. Program evaluation

1. A program evaluation was conducted to evaluate the effectiveness of the smoking cessation program. Substantiating data enhances the value of health promotion programs as key components of the health care delivery system. 2. Persons enrolled in the 8 week smoking cessation program and members of a comparison group were administered an evaluation tool prior to participating in the program, with a second evaluation tool upon completion of the program. Afterward, the smoking status of the members of both groups was evaluated at 3 month intervals for a period of 1 year. 3. Of the participants, 26.7% were not smoking 12 months after the program, compared to 6.9% of the comparison group. Of those participants who resumed smoking, 64% smoked less than half the amount they smoked before taking the class.     

Chromosomal response of insect and mammalian cells to streptonigrin: a comparative study

Respiratory syncytial virus (RSV) infections are characterized by upper or lower respiratory tract symptoms including bronchiolitis and pneumonia. Apnoea may be the first sign of disease in children with RSV infection. The aims of this study were the identification of independent risk factors for RSV associated apnoea and the prediction of the risk for mechanical ventilation in children with RSV associated apnoea. Medical records of children younger than 12 months of age admitted with RSV infection between 1992 and 1995 to the Sophia Children's Hospital, were reviewed. Demographic parameters, clinical features and laboratory parameters (SaO2, pCO2 and pH) were obtained upon admission and during hospitalization. Children with and without apnoea were compared using univariate and multivariate logistic and linear regression analysis. One hundred and eighty-five patients with RSV infection were admitted of whom 38 (21%) presented with apnoea. Patients with apnoea were significantly younger, had a significantly lower temperature, higher pCO2 and lower pH and had on chest radiographs also more signs of atelectasis. The number of patients admitted to the ICU because of mechanical ventilation and oxygen administration was significantly higher in children with RSV associated apnoea. Apnoea at admission was a strong predictor for recurrent apnoea. The relative risk for mechanical ventilation increased with the number of episodes of apnoea: 2.4 (95% CI 0.8-6.6) in children with one episode of apnoea (at admission) versus 6.5 (95% CI 3.3-12.9) in children with recurrent episodes of apnoea. CONCLUSIONS: Age below 2 months is the strongest independent risk factor for RSV associated apnoea. Apnoea at admission increases the risk for recurrent apnoea. The risk for mechanical ventilation significantly increases in children who suffer from recurrent apnoea.     

A method for the identification and quantitative investigation of denatured proteins in mixtures based on computer comparison of amino-acid patterns

Summary A procedure is described in which stepwise regression is adapted to permit comparison of the amino-acid pattern from a composite sample with those of simple substances arranged in an easily accessible data bank. The computer program automatically selects from the bank those proteins whose amino-acid patterns best correspond to that of the sample, and calculates the proportion of the proteins contained in the mixture.The procedure is applicable to food analytical problems that involve the identification and determination of proteins in mixtures, and also to situations in which the properties of the proteins have been altered by denaturation or enzymatic degradation. The determination is limited to 3–4 proteins in the mixture.

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Identifizierung und quantitative Bestimmung einzelner Proteine in Mischungen von denaturierten Proteinen mit Hilfe ihrer Aminosäauremuster

Zusammenfassung Mit Hilfe einer stufenweisen Regressionsberechnung konnen Aminosäu-renmuster eines zusammengesetzten Proteins mit einfachen Proteinen verglichen werden. Mit dieser Berechnung werden aus einer Sammlung von Vergleichsproteinen automatisch solche Proteine ausgewählt, deren Aminosäurenmuster am besten zu der der Probe paßt. Gleichzeitig können dadurch die Verhältnisse der verschiedenen Proteine in der Mischung berechnet werden. Durch these Arbeitsweise konnen weiterhin bei nahrungsanalytischen Problemen Proteine in Mischungen identifiziert and quantitativ erfaßt werden, selbst worn die Proteine denaturiert oder enzymatisch abgebaut sind, jedoch ist das Verfahren auf 3–4 Proteine in der Mischung begrenzt.

     

The electrical conductivities of poly(3.6-pyridazine) and poly(3.6-pyridazine sulphide)

Summary We have chemically synthesized poly(3.6-pyridazine),(PPd), and poly(3.6-pyridazine sulphide), (PPdS), and studied the electrical conductivities of these polymers.