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Biomedical subjects

C J Thompson

Publications and source records attributed to C J Thompson.

At least 19 recordsLinked to original sources
PubMed1992

Oxygen consumption of the living human brain measured after a single inhalation of positron emitting oxygen.

We measured the rate of washout of 15O-labeled water generated from labeled oxygen accumulated in brain after bolus [15O]O2 inhalation, and compared the washout with that of labeled water measured with H215O. Contrary to the original expectation, the radioactive water generated from labeled oxygen failed to leave the brain tissue at the rate predicted by exogenous water. Therefore, the use of a separately measured value for exogenous water clearance led to an error in the calculation of oxygen consumption. A new method presented in this paper eliminated the error by yielding oxygen consumption in a single oxygen study. We used time-weighted integration to estimate three parameters, including the unidirectional clearance from blood to brain (KO2(1)), the fractional clearance of the distribution volume in brain (kO2(2)), and the vascular volume correction (VO). We showed that the clearance of oxygen from blood to brain can be estimated with acceptable precision by this new approach, and that the new method yields a reliable measure of oxygen consumption.

Administration, Inhalation↗
PubMed1992

PETSIM: Monte Carlo simulation of all sensitivity and resolution parameters of cylindrical positron imaging systems.

Monte Carlo simulation techniques are applied to track the annihilation photons from positron decay, and store the photon histories. Reasonably realistic models of the isotope distribution in the brain and heart during typical PET studies, as well as the traditional phantoms used for measuring PET scanner performance can be built out of up to 10 hollow or solid cylinders. Separate programs model the source distribution and its attenuation characteristics, the collimators and the detectors. These modules are connected by compact gamma history files which are stored on disc or tape. Over 50 million gamma ray histories can be saved on a 1 Gbyte disc, representing the decay of several billion atoms. This allows for good precision even for single thin slices in scanners with wide axial acceptance. The simulation results include spectrum analysis, sensitivity to true coincident events, scattered coincident and single rays, and the effects on these parameters of detector dead time. The storage of intermediate results on tape reduces simulation time, since most common source geometries need be generated only once. The sensitivities in multi-slice systems are presented as matrices of coincident crystal planes. The matrix shows the true count sensitivity and the scatter fraction together for each valid combination of planes. This presentation is very useful for assessing the effects of various degrees of inter-plane collimation. The spatial resolution analysis includes the effects of positron range, non-collinearity of the gamma rays, multiple interaction within the detectors, and the effects of quantization into single crystals in multiple-crystal block detectors. Each of these effects can be turned on or off without repeating the simulation. Both in-plane and axial resolutions are calculated as a function of location of the positron-emitting nucleus and the angle of incidence of gamma rays on the crystals. Single crystals, blocks and crystals with depth of interaction encoding can be specified, as can the method of backprojection (planar, or 3D), so that the detector geometry can be optimized.

Computer Simulation↗
PubMed1992

The effects of the specific serotonin antagonist ICI 169,369 on the pituitary hormone response to insulin-induced hypoglycaemia in humans.

OBJECTIVE: To examine the role of serotonin in pituitary hormone release by studying the effect of a specific 5HT2 receptor antagonist, ICI 169,369, on the ACTH, prolactin, growth hormone and AVP response to insulin-induced hypoglycaemia in healthy humans. DESIGN: A double-blind, within-subject trial using a crossover design to compare the effect of placebo with two doses of ICI 169,369 on pituitary hormone responses to insulin-induced hypoglycaemia. PATIENTS: Ten healthy subjects were studied in the low-dose (30 mg x 2) limb and 11 healthy volunteers in the high-dose (80 mg x 2) limb. MEASUREMENTS: Plasma concentrations of prolactin, growth hormone, ACTH, cortisol and AVP, and blood glucose. RESULTS: In the low-dose study, pretreatment with 30 mg ICI 169,369, 10 and 2 hours before the study, had no effect on the fall in blood glucose or the rise in plasma ACTH, prolactin, growth hormone, AVP or plasma cortisol following insulin injection, when compared with placebo. In the high-dose study the effect of a higher dose (80 mg) of ICI 169,369 on the pituitary hormone response to hypoglycaemia was compared with that of placebo. Although the fall in blood glucose was similar following drug (4.3 +/- 0.1 to 1.5 +/- 0.5 mmol/l, mean +/- SEM, P less than 0.001) and placebo (4.3 +/- 0.1 to 1.4 +/- 0.4 mmol/l, P less than 0.001), the rise in plasma AVP was lower (P less than 0.05) following pretreatment with drug (0.5 +/- 0.2 to 2.1 +/- 0.6 pmol/l, P less than 0.05) than with placebo (0.7 +/- 0.2 to 3.4 +/- 0.9 pmol/l, P less than 0.01). CONCLUSIONS: The ACTH, prolactin, growth hormone and cortisol responses were unaffected by ICI 169,369. The data are compatible with an inhibitory effect of the serotonin antagonist ICI 169,369 on the AVP, but not the ACTH, prolactin or growth hormone response to insulin-induced hypoglycaemia in humans.

Adrenocorticotropic Hormone↗
PubMed1992

Global changes in gene expression related to antibiotic synthesis in Streptomyces hygroscopicus.

Two-dimensional gel electrophoresis was used to follow changes in gene expression associated with antibiotic (bialaphos) biosynthesis in Streptomyces hygroscopicus. Cultures were pulse-labelled with [35S]-methionine before, during, and after the switch from primary to secondary metabolism in order to compare kinetic profiles of bialaphos (antibiotic) production (bap) genes during this metabolic transition. Separation of gene products on two-dimensional gels revealed that 27 were dependent on brpA for optimal expression and were activated as the culture approached stationary phase. Genes which encoded 10 brpA-dependent proteins were mapped to a 10 kb SstI fragment of the 35 kb bap gene cluster by expressing them in Streptomyces lividans using the thiostrepton-inducible tipA promoter. N-terminal amino acid sequences of two brpA-dependent proteins, obtained by direct microsequencing of protein spots excised from two-dimensional gels, identified them as gene products mapping to the same region and involved in secondary metabolic conversions of the bap pathway. The kinetics of synthesis of 16 brpA-dependent gene products were characterized using QUEST computer software. Cluster analysis performed on the kinetics of synthesis of 346 of the most highly expressed gene products of HP5-29, including 16 brpA-dependent ones, identified 75 families having distinct patterns of expression. Many brpA-dependent proteins were clustered together; 10 were found in one kinetic family. These kinetic families also included brpA-independent gene products perhaps subject to similar regulatory mechanisms and thus possibly involved in bialaphos biosynthesis. The activation/derepression of bap expression took place as cultures approached stationary phase and was temporally related to synthesis of ppGpp.

Amino Acid Sequence↗
PubMed1992

Effects of human insulin on insulin binding antibody production in nondiabetic subjects.

OBJECTIVE: To test the hypothesis that human insulin may have a low immunogenicity and that short-term exposure may not cause endogenous insulin antibody production. RESEARCH DESIGN AND METHODS: Randomized double-blind prospective study. Serum samples collected for insulin binding antibodies and measured by a sensitive immunochemical assay. Subjects were seven healthy nondiabetic patients who had never received exogenous insulin. Each subject received 6 separate monthly injections of human insulin. On four occasions, both regular and NPH insulin were administered. On the other two occasions, either NPH or regular insulin was administered alone. RESULTS: Mean +/- SE basal insulin antibody levels (1.2 +/- 0.2 micrograms/L) increased to a maximal level of 4.5 +/- 0.8 micrograms/L after four injections. Thereafter, antibody levels declined to an end-of-study value of 2.5 +/- 0.3 micrograms/L. This represented a highly significant overall increase (P less than 0.001). A control group of six insulin-dependent diabetic subjects treated with human insulin over the same period as the test subjects demonstrated no change in insulin antibody concentrations (2.8 +/- 0.7-2.7 +/- 0.6 micrograms/L). CONCLUSIONS: These results suggest that human insulin preparations, when administered subcutaneously, may be more immunogenic than previously considered. The antigenic response was rapid, because only four subcutaneous injections were sufficient to produce insulin antibody levels in nondiabetic patients similar to those observed in insulin-dependent diabetic patients receiving chronic insulin replacement therapy.

Adult↗
PubMed1991

Regulation of ANP secretion in insulin-dependent diabetes mellitus and the influence of autonomic neuropathy.

In order to determine the effect of diabetic autonomic neuropathy (DAN) on the atrial natriuretic peptide (ANP) response to dynamic stimuli, we studied the ANP response to 60 degrees head-up and 60 degrees leg-up tilt in diabetic subjects with (DAN + ve, n = 8) and without (DAN - ve, n = 8) evidence of autonomic neuropathy and seven matched non-diabetic controls. Mean baseline plasma ANP concentrations were similar in all three groups. Head-up tilt was associated with a fall in plasma ANP in all seven healthy controls (21.8 (16.8-30.7) to 16.8 (7.1-29.1), P = 0.06, mean (range)), seven of the eight DAN - ve (16.9 (6.5-33.7) to 8.5 (3.0-21.1), P = 0.015) and all eight DAN + ve subjects (27.3 (8.5-101.5) to 15.4 (1.0-67.6), P = 0.044). Leg-up tilt caused a rise in plasma ANP in six of the seven healthy controls (17.6 (7.5-27.9) to 22.4 (15.2-48.1), P = 0.041), six of the eight DAN - ve (12.5 (7.8-27.8) to 15.5 (7.3-31.3), P = 0.054) and seven of the eight DAN + ve subjects (18.2 (2.8-55.1) to 25.1 (4.5-92.8), P = 0.013). There was no significant difference in the fall in plasma ANP during head-up tilt or in the rise in plasma ANP during leg-up tilt between the three groups. We conclude that the regulation of ANP secretion is normal in diabetes mellitus, and is unaffected by the presence of autonomic neuropathy.

Adult↗
PubMed1991

A comparison of the pharmacokinetics and metabolic effects of human regular and NPH insulin mixtures.

The effects of three human premixes (Mixtard, Actraphane, Humulin M3), syringe mixed 30% regular and 70% NPH insulin, regular insulin alone and NPH insulin alone, on intermediary metabolism, plasma free insulin levels and action profiles were compared using the euglycemic clamp technique. Seven normal volunteers received 20 IU of each insulin subcutaneously in a randomized fashion on separate days. The first and last 60 min of the 6 h clamp were chosen as summary measures of clinical importance. Significantly elevated plasma free insulin levels were found with all treatments compared to NPH insulin alone during the first hour, although by the final hour only Mixtard produced significantly higher levels compared to NPH (19.4 +/- 1.2, 10.5 +/- 0.3 mU/l P less than 0.01, respectively). Analysis of area under the incremental insulin absorption curve demonstrated that Mixtard produced significantly increased levels compared to syringe-mixed regular: NPH (7.6 +/- 0.8), Actraphane (9.6 +/- 1.0) and Humulin M3 (9.0 +/- 0.8 mU/l all P less than 0.05). Mixtard also resulted in significantly higher glucose infusion rates compared to the other premixes. No difference in action was found between regular and pre- or syringe-mixed human insulins during the first hour of the studies. The effects on intermediary carbohydrate and lipid metabolism were similar for syringe and premixed insulins. We conclude that: (1) fixed human insulin mixtures with NPH cause no blunting of the action of the soluble component. (2) Actraphane and Humulin M3 are similar but Mixtard may have a greater effect on some aspects of insulin action. (3) In clinical practice, fixed human insulin mixtures will be as efficacious as syringe-mixed preparations but may be easier and more convenient to use.

Adult↗
PubMed1991

Comparison of hospital and neighborhood controls in a study of coronary artery disease.

Case-control studies of risk factors for coronary artery disease (CAD) have almost invariably employed hospital controls, with minimal or no coronary artery stenosis. Although there is an important advantage in knowing the CAD status of controls, such groups are subject to bias related to hospitalization. To evaluate the generalizability of results obtained from studies using hospital controls, we compared risk factors in 342 hospital controls free of angiographic evidence for CAD, 168 neighborhood controls without symptoms of CAD, and 450 CAD patients. Coronary artery disease in cases and hospital controls was established arteriographically. No significant differences were found between the male control groups for total and low density lipoprotein (LDL) cholesterol, LDL apo-B, pack-years of smoking, body mass index, proportion with hypertension, diabetes and family history of coronary heart disease. Compared with neighborhood controls, male hospital controls had significantly lower high density lipoprotein (HDL) cholesterol, higher triglycerides and uric acid and scored higher on the Framingham Type A behavior pattern scale. Among women, the hospital control group had significantly lower LDL cholesterol and fewer pack-years of smoking, and a greater prevalence of hypertension than the neighborhood group. A greater proportion of both male and female hospital controls had left ventricular hypertrophy, and there were more current smokers among the neighborhood controls in both sexes. Age adjustment did not change these comparisons. While very few neighborhood controls were treated with beta-blockers, 32.7% of male and 41.4% of female hospital controls were so medicated. Control for beta-blocker use eliminated the difference in HDL cholesterol and triglycerides between the two male control groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Case-Control Studies↗
PubMed1991

The effects of collimation on PET image noise due to scatter, random counts, and deadtime.

The rates for true coincident events, scattered events, and singles in a Positron Emission Tomograph (PET) depend on the collimator size and shape. Monte Carlo techniques were used to compare efficiencies for wanted and unwanted events in multislice PET (MS-PET) and open collimator positron volumetric imaging (PVI) configurations. All systems used cylindrical arrays of bismuth germanate (BGO) or NaI crystals 51 cm in diameter and 10 cm deep suitable for whole brain imaging. The PVI systems detect about five times more true coincident events at low activity concentrations, but their scatter fraction is about three times higher. They are also much more sensitive to activity outside the scan field. As well as causing random counts when they fell within the energy acceptance window, single events are the main cause of deadtime. When the detectors are made from light-encoded blocks deadtime is the major limitation at high count rates. When discrete crystals are used, the efficiency is lower and the random count rates are a more significant source of noise. Noise-effective count rates are used to compare the relative cost in system performance among different systems and sources of noise.

Brain↗
PubMed1991

Osmotic and non-osmotic regulation of thirst and vasopressin secretion in patients with compulsive water drinking.

OBJECTIVE: To examine the osmotic and non-osmotic regulation of thirst and AVP release in patients with compulsive water drinking. DESIGN: A 2-hour intravenous infusion of hypertonic (855 mmol/l) sodium chloride solution, followed by a 2-hour drinking period. PATIENTS: Seven patients with compulsive water drinking, seven patients with diabetes insipidus and seven healthy controls. MEASUREMENTS: Plasma AVP, osmolality, sodium and haematocrit, thirst ratings on a visual analogue scale and the volume of water drunk in 2 hours following infusion. RESULTS: Plasma AVP responses to osmotic stimulation, and non-osmotic inhibition by drinking, were normal in patients with compulsive water drinking. Basal thirst ratings were higher in compulsive water drinking than in either diabetes (P less than 0.001) or controls (P less than 0.001), despite lower basal plasma osmolalities. There was a significant rise in thirst ratings during saline infusion, which correlated closely with plasma osmolality, in all three groups, but the final thirst ratings were higher in compulsive water drinkers, who subsequently drank more water than in either diabetes insipidus (P less than 0.01) or controls (P less than 0.001). Drinking rapidly lowered thirst ratings in controls and diabetes insipidus before changes occurred in plasma osmolality, but remained elevated in patients with compulsive water drinking. Linear regression analysis defined a lower osmotic threshold for thirst in compulsive water drinking compared with controls or diabetes insipidus. CONCLUSIONS: There are abnormalities of the osmotic stimulation and non-osmotic inhibition of thirst in compulsive water drinking, suggesting that the underlying defect is one of interpretation of osmotic and non-osmotic inputs. Measurement of thirst responses during hypertonic saline infusion and subsequent water drinking may provide useful diagnostic information in the differentiation of polyuric states.

Adult↗
PubMed1991

Site-specific integration of the Streptomyces plasmid pSAM2 in Mycobacterium smegmatis.

A method which allowed the stable integration of DNA fragments at a single site (attB) in the chromosome of Mycobacterium smegmatis was developed using an integrative element from Streptomyces ambofaciens, pSAM2. Vectors containing an Escherichia coli replicon (pBR322), the kanamycin resistance gene from Tn903 for selection in mycobacteria, and a fragment of pSAM2 containing the int gene as well as the attachment site (attP) were constructed and introduced to M. smegmatis by electroporation. Transformants showed stable integration of the plasmid into a single site (attB) of the mycobacterial genome. This approach should be valuable for analyses of gene expression in various mycobacterial species and permit the development of stable recombinant mycobacterial vaccine strains expressing bacterial or viral genes inserted in pSAM2.

Blotting, Southern↗
PubMed1991

Nucleotide sequence analysis reveals linked N-acetyl hydrolase, thioesterase, transport, and regulatory genes encoded by the bialaphos biosynthetic gene cluster of Streptomyces hygroscopicus.

Nucleotide sequence analysis of a 5,000-bp region of the bialaphos antibiotic production (bap) gene cluster defined five open reading frames (ORFs) which predicted structural genes in the order bah, ORF1, ORF2, and ORF3 followed by the regulatory gene, brpA (H. Anzai, T. Murakami, S. Imai, A. Satoh, K. Nagaoka, and C.J. Thompson, J. Bacteriol. 169:3482-3488, 1987). The four structural genes were translationally coupled and apparently cotranscribed from an undefined promoter(s) under the positive control of the brpA gene product. S1 mapping experiments indicated that brpA was transcribed by two promoters (brpAp1 and brpAp2) which initiate transcription 150 and 157 bp upstream of brp A within an intergenic region and at least one promoter further upstream within the bap gene cluster (brpAp3). All three transcripts were present at low levels during exponential growth and increased just before the stationary phase. The levels of the brpAp3 band continued to increase at the onset of stationary phase, whereas brpAp1-and brpAp2-protected fragments showed no further change. BrpA contained a possible helix-turn-helix motif at its C terminus which was similar to the C-terminal regulatory motif found in the receiver component of a family of two-component transcriptional activator proteins. This motif was not associated with the N-terminal domain conserved in other members of the family. The structural gene cluster sequenced began with bah, encoding a bialaphos acetylhydrolase which removes the N-acetyl group from bialaphos as one of the final steps in the biosynthetic pathway. The observation that Bah was similar to a rat and to a bacterial (Acinetobacter calcoaceticus) lipase probably reflects the fact that the ester bonds of triglycerides and the amide bond linking acetate to phosphinothricin are similar and hydrolysis is catalyzed by structurally related enzymes. This was followed by two regions encoding ORF1 and ORF2 which were similar to each other (48% nucleotide identity, 31% amino acid identity), as well as to GrsT, a protein encoded by a gene located adjacent to gramicidin S synthetase in Bacillus brevis, and to vertebrate (mallard duck and rat) thioesterases. The amino acid sequence and hydrophobicity profile of ORF3 indicated that it was related to a family of membrane transport proteins. It was strikingly similar to the citrate uptake protein encoded by the transposon Tn3411.

Amidohydrolases↗
PubMed1991

A survey of the heat shock response in four Streptomyces species reveals two groEL-like genes and three groEL-like proteins in Streptomyces albus.

A survey of the heat shock response was carried out in a series of streptomycetes. Four major heat shock proteins (HSPs) were observed in each of four species (Streptomyces albus, S. lividans, S. parvulus, S. viridochromogenes) after pulse labeling with [35S]methionine and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Three corresponded to the major procaryotic HSPs Lon, DnaK, and GroEL on the basis of their apparent molecular masses (94 to 100, 70, and 56 to 58 kDa, respectively). In addition, a smaller protein (16 to 18 kDa) was detected in all species but was most dramatically induced in S. albus. Consequently, studies focused on this species. As in other procaryotic systems, thermal induction (elicited by a shift from 30 degrees C to 41 degrees C) of the 70- and 94-kDa proteins was transient and expression returned to uninduced levels after 60 min. In contrast, the 56- to 58-kDa (GroEL) and 18-kDa proteins (HSP18) remained induced for more than 2 h. Two-dimensional gel electrophoresis allowed resolution of at least eight S. albus HSPs. HSP56-58 was composed of multiple acidic protein species, whereas HSP18 appeared to be basic. In spite of these differences in their physical characteristics, the N-terminal peptide sequence of HSP18 was similar to those of GroEL-like proteins found in other organisms and identical to one of the HSP56-58 species. In fact, N-terminal amino acid analysis of the S. albus 56- to 58-kDa species showed that it was composed of two proteins that differed in 3 of 10 positions, an observation that was supported by the detection of two groEL-like genes by Southern hybridization. The amino acid sequence of one of these proteins was identical to that of HSP18. Pulse-chase experiments did not reveal evidence of posttranslational processing of either HSP56-58 or HSP18.

Amino Acid Sequence↗
PubMed1991

Characterization of the groEL-like genes in Streptomyces albus.

Three GroEL-like heat shock proteins (HSP56, HSP58, and HSP18) have been observed in Streptomyces albus (G. Guglielmi, P. Mazodier, C. J. Thompson, and J. Davies, J. Bacteriol. 173:7374-7381, 1991). Here we report the cloning and complete nucleotide sequence of groEL1, which encodes HSP18 and HSP58, and groEL2, which encodes HSP56. Both nucleotide sequences predicted proteins of 56,680 Da that were 70% identical. The 5' nucleotide sequence of groEL1 coded for a protein corresponding to HSP18 that may be a processed gene product. At least two groEL-like genes were present in all 12 Streptomyces species tested; they were not closely linked in the genome. groEL1, but not groEL2, was adjacent to a groES-like gene.

Amino Acid Sequence↗
PubMed1991

Reproducibility of osmotic and nonosmotic tests of vasopressin secretion in men.

We have studied the reproducibility of the thirst and arginine vasopressin (AVP) responses to osmotic and hypoglycemic stimulation in healthy volunteers undergoing repeat hypertonic (855 mmol/l) saline infusion and insulin tolerance tests (ITTs). Hypertonic saline infusion caused similar mean rises in plasma osmolality, AVP, and thirst on each occasion. Linear-regression analysis defined close relationships between the slopes (r = +0.72, P less than 0.05) and the abscissal intercepts (r = +0.89, P less than 0.001) of the regression lines relating plasma osmolality (Posmol) and plasma AVP (PAVP), and the group intraindividual component of the variance for the slopes and intercepts was 7 and 0.6%, respectively. There were close correlations between the slopes (r = +0.79, P less than 0.02) and the intercepts (r = +0.84, P less than 0.01) of the regression lines relating Posmol and thirst, and group intraindividual component of the variance was 14 and 0.7%, respectively. Hypertonic saline infusion was infused on four occasions in four subjects, and the results showed that the linear regression lines relating PAVP and Posmol and thirst and Posmol were reproducible within an individual. There were similar falls in blood glucose and elevations in PAVP in both ITTs. No relationship was defined between the fall in blood glucose and either the rise in PAVP or the area under the AVP curve (AUC). The intraindividual component of the variance for the rise in AVP and the AUC was 77 and 22.5%, respectively. The AVP and thirst responses to osmotic stimulation are highly reproducible, but there is considerable intraindividual variation in the AVP response to hypoglycemia.

Adult↗
PubMed1991

Validation of simultaneous PET emission and transmission scans.

A technique for performing simultaneous PET emission and transmission scans is validated in a fluoro-deoxyglucose study. A point source masked into a fan beam of annihilation photons orbits the patient section under study. Coincident events are sorted into two buffers, or rejected, based on the source's position. Both static and dynamic frames of independent and simultaneous studies are compared. The noise effective count rate is reduced to 62% of the value during normal studies. However, the increase in the coefficient of variation in cortical regions is less than 6%. The RMS difference between profile contours through many brain regions is approximately 40% higher comparing two simultaneous emission/transmission scans than when the same analysis is performed on independent emission scans. This difference appears to be due to the noise patterns arising from the use of different transmission scans.

Adult↗
PubMed1990

Streptomyces hygroscopicus has two glutamine synthetase genes.

Streptomyces hygroscopicus, which produces the glutamine synthetase inhibitor phosphinothricin, possesses at least two genes (glnA and glnB) encoding distinct glutamine synthetase isoforms (GSI and GSII). The glnB gene was cloned from S. hygroscopicus DNA by complementation in an Escherichia coli glutamine auxotrophic mutant (glnA). glnB was subcloned in Streptomyces plasmids by insertion into pIJ486 (pMSG3) and pIJ702 (pMSG5). Both constructions conferred resistance to the tripeptide form of phosphinothricin (bialaphos) and were able to complement a glutamine auxotrophic marker in S. coelicolor. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of S. lividans(pMSG5) revealed a highly overexpressed 40-kilodalton protein. When GS was purified from this strain, it was indistinguishable in apparent molecular mass from the 40-kilodalton protein. The nucleic acid sequence of the cloned region contained an open reading frame which encoded a protein whose size, amino acid composition, and N-terminal sequence corresponded to those of the purified GS. glnB had a high G + C content and codon usage typical of streptomycete genes. A comparison of its predicted amino acid sequence with the protein data bases revealed that it encoded a GSII-type enzyme which had previously been found only in various eucaryotes (47 to 50% identity) and nodulating bacteria such as Bradyrhizobium spp. (42% identity). glnB had only 13 to 18% identity with eubacterial GSI enzymes. Southern blot hybridization experiments showed that sequences similar to glnB were present in all of the five other Streptomyces species tested, as well as Frankia species. These results do not support the previous suggestion that GSII-type enzymes found in members of the family Rhizobiaceae represent a unique example of interkingdom gene transfer associated with symbiosis in the nodule. Instead they imply that the presence of more than one gene encoding GS may be more common among soil microorganisms than previously appreciated.

Amino Acid Sequence↗