Biomedical subjects
Publications and source records attributed to J Kingma.
The incidence of accidental fall in 67,436 trauma patients was investigated for the period 1971-1995. Over-all a decrease in incidence was found from 1976 through 1995; however, notwithstanding the over-all picture a strong age-specific increase in the incidence was observed in persons of 80 years and over from 1981-1985 through 1991-1995.
Haloalkane dehalogenase converts haloalkanes to their corresponding alcohols. The 3D structure, reaction mechanism and kinetic mechanism have been studied. The steady state k(cat) with 1,2-dichloroethane and 1,2-dibromoethane is limited mainly by the rate of release of the halide ion from the buried active-site cavity. During catalysis, the halogen that is cleaved off (Cl alpha) from 1,2-dichloroethane interacts with Trp125 and the Cl beta interacts with Phe172. Both these residues have van der Waals contacts with Val226. To establish the effect of these interactions on catalysis, and in an attempt to change enzyme activity without directly mutating residues involved in catalysis, we mutated Val226 to Gly, Ala and Leu. The Val226Ala and Val226Leu mutants had a 2.5-fold higher catalytic rate for 1,2-dibromoethane than the wild-type enzyme. A pre-steady state kinetic analysis of the Val226Ala mutant enzyme showed that the increase in k(cat) could be attributed to an increase in the rate of a conformational change that precedes halide release, causing a faster overall rate of halide dissociation. The k(cat) for 1,2-dichloroethane conversion was not elevated, although the rate of chloride release was also faster than in the wild-type enzyme. This was caused by a 3-fold decrease in the rate of formation of the alkyl-enzyme intermediate for 1,2-dichloroethane. Val226 seems to contribute to leaving group (Cl alpha or Br alpha) stabilization via Trp125, and can influence halide release and substrate binding via an interaction with Phe172. These studies indicate that wild-type haloalkane dehalogenase is optimized for 1,2-dichloroethane, although 1,2-dibromoethane is a better substrate.
OBJECTIVE: To assess the short-term and long-term outcomes of pediatric polytrauma patients and to analyze the extent to which short-term outcomes can predict long-term outcomes. MATERIALS AND METHODS: All pediatric polytrauma patients (Injury Severity Score of > or = 16, < or = 15 years of age) who were treated at a Level I trauma center between 1985 and 1989 and who were 18 years or older at follow-up (1996) were included. Short-term outcomes were quantified using the Glasgow Outcome Scale. The Functional Independence Measure, the RAND-36, and questions on social outcome were used to assess long-term outcomes. MEASUREMENTS AND MAIN RESULTS: Fifty-nine of 74 patients survived (80%). At 1 year after injury, 22% were disabled, mainly due to severe brain injuries, and 32% had changed school. After 9 years of follow-up, the degree of physical disablement was low (12%). Cognitive impairments were found in 42% of the patients. Only 10% were in receipt of disablement allowances; 76% were employed or attended school. The quality of life enjoyed by the patients (RAND-36) did not differ from that enjoyed by a healthy reference population. The Glasgow Outcome Scale (from 6 weeks onward) was a good predictor of long-term disablement. CONCLUSIONS: Although the short-term impact of injuries is extensive, the long-term outcomes are satisfactory. Severe injuries sustained during childhood do not lead to a diminished quality of life during adulthood.
This retrospective study concerns the aetiology and psychological long-term effects of injuries due to bicycle accidents in 329 patients 50 years and older who attended the Emergency Unit of the University Hospital at Groningen during the period 1990 through 1992. Long-term effects were assessed three years after hospital discharge. The one-sided bicycle accident (with no other traffic involved) was the major (63.2%) cause, mostly due to loss of balance or to a foot slipping from the pedal. The main category of the second major cause was collision with other traffic. Of the bicycle accidents 66% occurred within 15 minutes after departure; 80% of the accidents happened in good weather conditions and daylight, and 7.6% of the patients had taken tranquilizers before biking. The majority of the injuries were observed at the upper extremities (28.8%) and head or face (25.8%). The percentage of clinically treated patients increased across ages from 25% in the 50- to 54-yr.-old category to 45% in the category 75-yr. and older. Three years after the incident, long-term psychological effects were still observed in 29% of the patients.
One therapeutic alternative for redislocation of Colles' fractures is closed reduction and transstyloid Kirschner-wire fixation. We describe our results concerning 21 redislocations treated in this way. According to Older's classification, B fractures were type 3 and 13 fractures type 4. After a median follow-up period of 2 years, most patients had regained normal volar tilt, but significant loss of radial tilt and radial length was found in 11 patients. Malunion occurred in 8 wrists due to either fracture comminution or insufficient K-wire fixation. According to Gartland and Werley's point system, the end-results were poor in 2, fair in 11, good in 4, and excellent in 4 wrists. There was no statistically significant difference in secondary displacement and functional end-result between Older's type 3 and type 4 fractures. We conclude that K-wire fixation is frequently not firm enough and external fixation might be a better alternative for stabilizing redislocated distal radial fractures.
Haloalkane dehalogenase converts halogenated alkanes to their corresponding alcohols. The active site is buried inside the protein and lined with hydrophobic residues. The reaction proceeds via a covalent substrate-enzyme complex. This paper describes a steady-state and pre-steady-state kinetic analysis of the conversion of a number of substrates of the dehalogenase. The kinetic mechanism for the "natural" substrate 1,2-dichloroethane and for the brominated analog and nematocide 1,2-dibromoethane are given. In general, brominated substrates had a lower Km, but a similar kcat than the chlorinated analogs. The rate of C-Br bond cleavage was higher than the rate of C-Cl bond cleavage, which is in agreement with the leaving group abilities of these halogens. The lower Km for brominated compounds therefore originates both from the higher rate of C-Br bond cleavage and from a lower Ks for bromo-compounds. However, the rate-determining step in the conversion (kcat) of 1, 2-dibromoethane and 1,2-dichloroethane was found to be release of the charged halide ion out of the active site cavity, explaining the different Km but similar kcat values for these compounds. The study provides a basis for the analysis of rate-determining steps in the hydrolysis of various environmentally important substrates.
Pseudomonas sp. GJ1 is able to grow with 2-chloroethanol as the sole carbon and energy source, but not with 2-bromoethanol, which is toxic at low concentrations (1 mM). A mutant that could grow on 2-bromoethanol with a growth rate of 0.034 h-1 at concentrations up to 5 mM was isolated and designated strain GJ1M9. Measurement of enzyme activities showed that mutant and wild-type strains contained a PMS-linked alcohol dehydrogenase that was active with halogenated alcohols and that was threefold overexpressed in the mutant when grown on 2-chloroethanol, but only slightly overproduced when grown on 2-bromoethanol. Both strains also contained an NAD-dependent alcohol dehydrogenase that had no activity with halogenated alcohols. Haloacetate dehalogenase levels were similar in the wild-type and the mutant. Activities of NAD-dependent aldehyde dehydrogenase were only slightly higher in extracts of the mutant grown with 2-bromoethanol than in those of the wild-type grown with 2-chloroethanol. SDS-PAGE, however, showed that this enzyme amounted to more than 50% of the total cellular protein in extracts of the mutant from 2-bromoethanol-grown cells, which was fourfold higher than in extracts of the wild-type strain grown on 2-chloroethanol. The enzyme was purified and shown to be a tetrameric protein consisting of subunits of 55 kDa. The enzyme had low Km values for acetaldehyde and other non-halogenated aldehydes (0.8-4 microM), but much higher Km values for chloroacetaldehyde (1.7 mM) and bromoacetaldehyde (10.5 mM), while V(max) values were similar for halogenated and non-halogenated aldehydes. Cultures that were pregrown on 2-chloroethanol rapidly lost aldehyde dehydrogenase activity after addition of 2-bromoethanol and chloroamphenicol, which indicates that bromoacetaldehyde inactivates the enzyme. To achieve growth with 2-bromoethanol, the high expression of the enzyme thus appears to be necessary in order to compensate for the high Km for bromoacetaldehyde and for inactivation of the enzyme of bromoacetaldehyde.
Haloalkane dehalogenase (DhlA) from Xanthobacter autotrophicus GJ10 catalyzes the hydrolytic cleavage of carbon-halogen bonds in a broad range of halogenated aliphatic compounds. Previous work has shown that Asp124, which is located close to the internal substrate-binding cavity, carries out a nucleophilic attack on the C-alpha of the alkylhalide, displacing the halogen. The resulting alkyl-enzyme intermediate is subsequently hydrolyzed. In order to study the role of His289 in the hydrolysis of the intermediate, a His289-->Gln mutant was constructed by site-directed mutagenesis. The purified mutant enzyme was not catalytically active with haloalkanes, but a halide burst stoichiometric to the amount of enzyme was observed with 1,2-dibromoethane. Using ion spray mass spectrometry, accumulation of the covalent alkyl-enzyme and binding of the alkyl moiety of the substrate to an Asp124-containing tryptic peptide were shown. Fluorescence-quenching experiments indicated that halide ions are strongly bound by the alkyl-enzyme but not by the substrate-free enzyme. The results show that His289 is the base catalyst for the dealkylation of the covalent intermediate, but that it is not essential for the initial nucleophilic attack of Asp124 on the C-1 atom of the haloalkane. Furthermore, the halide ion that is released in the first step probably leaves the active site only after hydrolysis of the alkyl-enzyme.
Haloalkane dehalogenase catalyzes the hydrolytic cleavage of carbon-halogen bonds in short-chain haloalkanes. Two tryptophan residues of the enzyme (Trp125 and Trp175) form a halide-binding site in the active-site cavity, and were proposed to play a role in catalysis. The function of these residues was studied by replacing Trp125 with phenylalanine, glutamine or arginine and Trp175 by glutamine using site-directed mutagenesis. All mutants except Trp125-->Phe showed a more than 10-fold reduced kcat and much higher Km values with 1,2-dichloroethane and 1,2-dibromoethane than the wild-type enzyme. Fluorescence quenching experiments showed a decrease in the affinity of the mutant enzymes for halide ions. The 2H kinetic isotope effect observed with the wild-type enzyme in deuterium oxide was lost in the active mutants, except the Trp125-->Phe enzyme. The results indicate that both tryptophans are involved in stabilizing the transition state during the nucleophilic substitution reaction that causes carbon-halogen bond cleavage.
Haloalkane dehalogenase hydrolyses various 1-halo-n-alkanes to the corresponding alcohols by covalent catalysis with formation of an alkyl-enzyme intermediate. The carboxylate function of the nucleophilic aspartate (Asp-124) that displaces the halogen during formation of the intermediate was changed to an amide by site-directed mutagenesis (Asp-124-->Asn). Activity measurements and analysis of peptides containing the nucleophilic residue showed that the mutant enzyme was inactive, but that the activity increased by rapid deamidation of the asparagine residue, yielding wild type enzyme. There was no indication for isoaspartate formation during this process. The results suggest that a water molecule that is located close to the carboxyl function of Asp-124 in the X-ray structure is highly reactive and is responsible for the observed deamidation.
Haloacid dehalogenases are enzymes that cleave carbon-chlorine or carbon-bromine bonds of 2-haloalkanoates. X-ray-quality crystals of L-2-haloacid dehalogenase from the 1,2-dichloroethane-degrading bacterium Xanthobacter autotrophicus GJ10 have been grown at room temperature from 20% PEG 8000, 200 mM sodium formate at pH 6.8-7.0, using macroseeding techniques. The crystals, which diffract in the X-ray beam up to 2.0 A resolution, belong to the spacegroup C2221. Cell parameters are a = 58.8 A, b = 93.1 A, c = 84.2 A. A native data set to 2.3 A has been collected, with a completeness of 97% and an Rsym of 6.0%.
The present purpose was to illustrate the range of severity scores among trauma patients. A computerized conversion of ICD-9CM injury diagnoses into scores for severity of injury was processed on data of 36,178 trauma patients. More than 94% of the patients had injuries of minor severity. The probability of mortality was .67 when the Injury Severity Score (ISS) was > or = 16.
Haloalkane dehalogenase catalyzes the hydrolytic cleavage of carbon-halogen bonds in a broad range of halogenated aliphatic compounds. The X-ray structure suggests that Asp124, which is located close to an internal cavity, carries out a nucleophilic attack on the C alpha of the substrate, releasing the halogen. To study the mechanism of hydrolysis, this aspartate residue was mutated to alanine, glycine, or glutamate. The mutant enzymes showed no activity toward 1,2-dichloroethane and 1,2-dibromoethane. Incubation of purified wild-type dehalogenase with 1,2-dichloroethane in the presence of H2(18)O resulted in the incorporation of 18O in 2-chloroethanol and in the carboxylate group of Asp124. This shows that the reaction proceeds by covalent catalysis with the formation of an alkyl-enzyme intermediate that is hydrolyzed by attack of solvent water on the carbonyl carbon of Asp124. On the basis of amino acid sequence similarity between haloalkane dehalogenase and epoxide hydrolases, it is proposed that a conserved aspartate residue is also involved in covalent catalysis by the latter enzymes.
In this study, we review the incidence and aetiology of 3858 metacarpal fractures in a series of 235,427 patients across the life-span. We found men aged 10-29 to have the highest incidence rates for metacarpal fracture (2.5 per cent). Accidental fall was the dominant cause of these fractures in the age group 9 years or less (38.7 per cent) and in the groups 50 years and older. Our data clearly show transport accidents and in particular bicycle accidents to be major determinants in all age groups. We conclude that recognition of the high frequency of metacarpal fractures among men aged 10-29 years should be given and public expenditure should be invested in the development and recommendation of proper safety gear.
The incidence and aetiology of 6,857 phalangeal fractures of the hand have been reviewed in a series of 235,427 patients, looking for an age-specific vulnerability to fracture. We found sports to be the main cause of fracture in the 10-29 years age groups and accidental falls to be the leading cause in those aged 70 years or older. We made a new observation that the highest incidence occurs in the male 40-69 age group and machinery was the dominant cause of fracture in this group. Recognition of the frequency of industrial trauma is needed, and public expenditure should be invested in its prevention and treatment.
Diagnoses of injuries as a result of trauma are commonly coded by means of the International Classification of Diseases (9th rev.) Clinical Modification (ICD-9CM). The Abbreviated Injury Scale (AIS) is frequently employed to assess the severity of injury per body region. The Injury Severity Score (ISS) is an over-all index or summary of the severity of injury. To compute one of these two types of scores the entire medical record of each patient must be examined. The program ICDTOAIS replaces the manual coding or translation between the two scores. The program converts the ICD-9CM coded diagnoses into AIS and ISS scores. The program also computes the maximum AIS (MAXAIS) per body region, enabling the researcher to assess the relative impact of the severity of trauma of different body regions in both morbidity and mortality studies. The program locates invalid ICD-9CM rubrics in the data file. ICDTOAIS may be employed as a program alone or as a procedure in database management systems (e.g., DBase III plus, DBase IV, or the different versions of FOXPRO). The program is written in Turbo Pascal, Version 6.
SHOWICD is an interactive computer program designed to document severity of injury from the ICD-9CM coded injury diagnoses of a particular patient. Two severity-of-injury scores [the Abbreviated Injury Scale (AIS) and the Injury Severity Score (ISS)] are used. By employing the AIS scores, the severity of injury may be assessed per body region. The ISS provides an over-all index or summary score for severity of injury of the whole body. SHOWICD allows the user to analyze the effects of different types of injuries on the Injury Severity Score. SHOWICD may be employed either alone as a program or as a procedure in database management systems. The program is written in Turbo Pascal.
The causes of occupational injuries (N = 2,365) were investigated. Accidents with machinery and hand tools were the two main causes (49.9%). 89% of the patients with occupational injuries were male. The highest risk group were in the age category of 19 years or less (51.9%). This age group also showed the highest incidence rate for females.