The temperature-sensitive bond in human alpha 2-macroglobulin is the alkylamine-reactive site.
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Amines↗
Biomedical subjects
J B Howard
Publications and source records attributed to J B Howard.
Comparison of the iron proteins from the nitrogen fixation complexes of Azotobacter vinelandii, Clostridium pasteurianum, and Klebsiella pneumoniae.
The molecular weights, amino acid compositions, amino- and carboxyl-terminal sequences, and ion-exchange peptide maps of the cysteine-containing tryptic peptides were determined for the iron proteins from the nitrogen fixation complexes of Azotobacter vinelandii (Av2) and Klebsiella pneumoniae (Kp2). Our results are compared to the known amino acid sequence of the iron protein from Clostridium pasteurianum (Cp2) [Tanaka, M., Haniu, M., Yasunobu, K. & Mortenson, L. E. (1977) J. Biol. Chem. 252, 7093-7100]. Previous studies have shown the iron proteins to have similar enzymatic functions and spectroscopic properties. Furthermore, the DNAs coding for the iron protein from many different species cross-hybridize [Ruvkun, G. B. & Ausubel, F. M. (1980) Proc. Natl. Acad. Sci. USA 77, 191-195]. Our results indicate that the protein structures are similar yet have significant differences. The amino-terminal sequences of Av2 and Kp2 are extended compared to the amino-terminal methionine of Cp2 and may indicate a different initiation site in these proteins. The aminoterminal sequences for Av2 and Kp2 are more homologous with each other than either of these are with Cp2. The carboxyl-terminal sequences are extended in Av2(14 residues) and Kp2 ( approximately 30 residues) compared to Cp2. The amino- and carboxyl-terminal sequences establish that either the structural gene sizes are different in the three organisms or extensive posttranslational modification must occur in some species. Because cysteinyl residues are involved at the active site of the iron protein, a sensitive peptide mapping technique was used to compare cysteinyl peptides of the iron protein from the three species. Av2 and Kp2 have a redistribution of cysteinyl residues when compared to Cp2. Three important differences in the cysteine distributions were found, namely, residue 4 is valine and residue 148 is alanine in Cp2, but cysteinyl residues occupy these positions in Av2, whereas residue 231 is cysteine in Cp2 but alanine in Av2. The peptide mapping technique provides a method for the investigation of selective chemical modification of cysteinyl residues.
Amino Acid Sequence↗
The primary structure of the alpha subunit of protocatechuate 3,4-dioxygenase. I. Isolation and sequence of the tryptic peptides.
The carboxymethylated alpha subunit of protocatechuate 3,4-dioxygenase was digested with trypsin. The 14 tryptic peptides were isolated by ion exchange chromatography on DEAE-Sephadex and by gel filtration chromatography. Automated Edman degradation and carboxypeptidase Y and B digestion were used to establish the sequence of these peptides. Further fragmentation of two tryptic peptides, T3 and T5, by Staphylococcus aureus protease and cyanogen bromide, respectively, was necessary to complete the sequences. The tryptic peptides accounted for a minimum of 199 residues out of a total of 202 residues predicted by amino acid analysis.
Amino Acid Sequence↗
The primary structure of the alpha subunit of protocatechuate 3,4-dioxygenase. II. Isolation and sequence of overlap peptides and complete sequence.
The complete primary structure of the alpha subunit of protocatechuate 3,4-dioxygenase has been determined by automated Edman degradation and carboxypeptidase digestionof the intact alpha chain and of peptides derived from trypsin (N.A. Kohlmiller and J.B. Howard (1979) J. Biol. Chem. 254, 7302-7308) and Staphylococcus aureus protease digestion, and from hydroxylamine and dilute acid cleavage. The alpha chain was found to consist of 200 residues in the following sequence from the NH2-terminal end: (formula: see text).
Amino Acid Sequence↗
Structural characterization of human alpha2-macroglobulin subunits.
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Amino Acid Sequence↗
Characterization of alkylamine-sensitive site in alpha 2-macroglobulin.
Methylamine reacts with the plasma protease inhibitor, alpha 2-macroglobulin, to form an irreversible, covalent modification. Quantitation of the reaction indicates 3.9 +/- (SD) 0.4 reactive sites per native tetrameric protein (Mr = 725,000) or one site per subunit. The reaction is selective and specific in that only 1 or 2 labeled peptides are observed on radioautography of peptide maps derived from [14C]methylamine-treated alpha 2-macroglobulin. A single chymotryptic peptide was isolated in 56% overall yield from the labeled protein. The peptide sequence by Edman degradation was found to be Gly-Cys-Gly-Glu-X-Asn-Met-(Val, Leu), in which X was the only radiolabeled phenylthiohydantoin derivative. Amino acid analysis and mass spectral analysis of the derivative suggests that X is gamma-glutamylmethylamide. Because glutamic acid and glutamine residues do not normally react with alkylamines, this work presents presumptive evidence for an alternative activated center in selected proteins.
Binding Sites↗
Report of 65 cases of Haemophilus influenzae b pneumonia.
During a 14-year period there were 65 cases of pneumonia caused by Haemophilus influenzae b; 83% were children less than 2 years of age and 80% of illness occurred in winter and spring. The roentgenographic picture was consolidative pneumonia in 75% and pleural effusions were present in 75% of all cases. Ten patients had associated meningitis and three had purulent pericarditis. Otitis media was diagnosed in 43% and H influenzae b was isolated from eight middle ear aspirates. Three patients (5%) died. Recommendations for diagnosis and treatment are made.
Anti-Bacterial Agents↗
Isolation and properties of the trypsin-derived ADP-ribosyl peptide from diphtheria toxin-modified yeast elongation factor 2.
We have developed a method for the purification in micromole amounts of the trypsin-derived ADP-ribosyl peptide from diphtheria toxin-modified yeast elongation factor 2 (EF-2). EF-2 was partially purified (15 to 20% purity) by ammonium sulfate precipitation and DEAE-Sephadex chromatography. After [3H]ADP-ribosylation by [3H]nad+ and diphtheria toxin, EF-2 was digested with trypsin and a homogeneous [3H]ADP-ribosyl peptide was isolated by chromatography on DEAE-Sephadex and dihydroxyboryl-substituted cellulose. Based on the amount of ADP-ribose acceptor activity in the crude extract, the overall yield of the peptide was 35%. The yeast peptide contains an unusual amino acid (X) which is the site of ADP ribosylation and is apparently identical to the amino acid reported from rat liver EF-2 by Robinson et al. (Robinson, E. A., Hendriksen, O., and Maxwell, E.S. (1974) J. Biol. Chem. 249, 5088-5093). The sequence of the 15-residue yeast peptide was determined to be: Val-Asn-Ile-Leu-Asp-Val-Thr-Leu-His-Ala-Asp-Ala-Ile-X-Arg. The 11 COOH-terminal residues of this peptide and of the homologous 15-residue peptide reported by Maxwell and co-workers from rat liver EF-2 are identical.
Adenosine Diphosphate Sugars↗
Isolation and partial characterization of two different subunits from the molybdenum-iron protein of Azotobacter vinelandii nitrogenase.
The molybdenum-iron protein of Azotobacter vinelandii nitrogenase was separated into two subunits of equal concentration by ion exchange chromatography on sulfopropyl (SP) Sephadex at pH 5.4 in 7 M urea. Better than 90% yield of each subunit was obtained on a preparative scale if the reduced carboxymethylated molybdenum-iron protein was incubated at 45 degrees C for 45 min prior to chromatography. Without the heating step low yields of the subunits were obtained. Although the amino acid compositions of the two subunits were very similar, the NH2-terminal sequences were completely different as determined by automated sequential Edman degradation. The sequence for the alpha subunit was NH2-Ser-Gln-Gln-Val-Asp-Lys-Ile-Lys-Ala-Ser-Tyr-Pro-Leu-Phe-Leu-Asp-Gln-Asp-Tyr- and for the beta subunit the sequence was NH2-Thr-Gly-Met-Ser-Arg-Glu-Glu-Val-Glu-Ser-Leu-Ile-Gln-Glu-Val-Leu-Glu-Val-Tyr-. Likewise the COOH-terminal sequences for the two subunits, as determined with carboxypeptidase Y, were tota-ly different. The sequence for the alpha subunit was -Leu-Arg-Val-COOH and that for the beta subunit was -Ile-(Phe, Glu)-Ala-Phe-COOH. Radioautographs of tryptic peptide maps were prepared for the molybdenum-iron protein and the two subunits which had been labeled at the cysteinyl residues with iodo[2-14C]acetic acid. These maps indicated that the two subunits had no cysteinyl peptides in common and that the cysteinyl residues were clustered in both subunits.
Amino Acid Sequence↗
Trimethoprim-sulfamethoxazole vs sulfamethoxazole for acute urinary tract infections in children.
A total of 118 children between 6 months and 10 years of age with acute urinary tract infection were treated in a random; double-blind manner with 12 mg/kg/day of trimethoprim-sulfamethoxazole (61 patients) or 50 mg/kg/day of sulfamethoxazole (57 patients) for ten days. Mean trimethoprim and sulfamethoxazole susceptibilities of Escherichia coli isolated from these patients were 1.2 and 0.6 microgram/ml, respectively. Mean serum concentrations of trimethoprim and sulfamethoxazole were 1.8 and 62 microgram/ml, respectively, one hour after the dose. Of the children who completed the ten days of prescribed medication, clinical and bacteriological cure was confirmed immediately after treatment for all but one patient in each group. Most patients in each treatment group with recurrent infections had underlying urological abnormalities. Severe hematological, renal, or hepatic toxicity requiring interruption of treatment was not encountered. No advantage of trimethoprim-sulfamethoxazole over sulfamethoxazole alone for acute urinary tract infection was demonstrated.
Acute Disease↗
Oral antibiotic therapy for skeletal infections of children. I. Antibiotic concentrations in suppurative synovial fluid.
To evaluate the feasibility of oral antibiotic treatment for pyogenic arthritis, one or more oral doses of antibiotics were substituted for the drugs being used for parenteral therapy. Synovial fluid and serum specimens obtained at randomized times after an oral dose of ampicillin, cephalexin, cloxacillin, dicloxacillin, or penicillin G were assayed for antibiotic content and antibacterial activity. Seventy specimens from 21 infants and children were studied. Peak synovial fluid concentrations were greater than 60% of peak serum concentrations with all drugs tested and there was adequate inhibitory activity against bacteria commonly causing arthritis. The degree of antibiotic binding to serum protein had no apparent effect on the degree of penetration into pyogenic synovial fluid.
Administration, Oral↗
Antibiotic concentrations in pus and bone of children with osteomyelitis.
Pharmacologic data are presented on 39 children treated for osteomyelitis with one of the following antibiotics: methicillin, dicloxacillin, cephaloridine, or cefazolin. The concentrations of drug in pus and bone were correlated with serum concentrations, with the susceptibilities of Staphylococcus aureus strains isolated from the patients, and with the degree of drug protein-binding. The penetration of the antibiotics into pus and bone was similar for the two penicillins and for the two cephalosporins despite the disparate protein-binding affinities of these drugs. The agents attained concentrations in tissues that were at least several fold, and often more than tenfold, greater than the MIC and MBC values of the S. aureus stains. These data provide a basis for selection of antimicrobial agents for treatment of osteomyelitis.
Adolescent↗
Nursing home medication costs.
The cost of discarded drugs at a level-III nursing home was calculated for a four-year period. A significant percentage of the cost seemed to be the result of poor prescribing practices. Prescriptions for antibiotics and for psychotropic drugs (particularly PRN orders) are discussed in some detail. Medication policies were developed to improve physicians' prescribing habits in order to reduce these costs without jeopardizing the quality of care.
Costs and Cost Analysis↗
Nurses' notes on nursing home patients: are they effective?
The notes written by nurses in a level-III nursing home were reviewed in detail. They were found to be neither pertinent nor meaningful in most cases. The daily writing of these notes required a large percentage of the nurses' time but they were seldom, if ever, read by physicians. An alternate system of writing notes on a weekly basis is suggested.
Costs and Cost Analysis↗
Source identification of urban airborne polycyclic aromatic hydrocarbons by gas chromatographic mass spectrometry and high resolution mass spectrometry.
Polycyclic aromatic hydrocarbons formed during the combustion of three common fuels (coal, wood and kerosene) were separated and identified by capillary-column gas chromatographic mass spectrometry and were compared to airborne polycyclic aromatic hydrocarbons from Indianapolis, a high coal consuming area, and Boston, a low coal consuming area. High resolution mass spectral data were utilized in the construction of alkyl homolog plots for the comparison of alkyl distribution within each sample.
Air Pollution↗
Medication procedures in a nursing home: abuse of PRN orders.
Nursing home patients often receive more drugs than needed. Drug incompatibilities frequently are found, and PRN orders are apt to be overused. Regulatory agencies require periodic physician review in the hope of eliminating these loose prescribing habits. A study was made of the drugs prescribed for 98 patients in a proprietary nursing home. The principal findings were that PRN orders can indeed be inappropriate and indiscriminately followed, and that the multiplicity of drugs often prescribed can lead to drug incompatibilities and to additional illness of the patient. Concern is expressed about the medication review process currently in vogue in nursing homes.
Aged↗
Evaluating the quality of nursing home care.
Evaluation of the quality of care that a nursing home patient receives is difficult and requires physician participation as well as the participation of specially trained nursing personnel. A form of limited medical audit is suggested to aid in such evaluation. In the proprietary nursing home studied by the authors, the quality of care was not being effectively assessed by the survey process then employed. In addition to other factors, emphasis is now placed on the avoidance of additional costs or extra paper work in the process of determining the quality of care.
Aged↗
Residual effects of septic arthritis in infancy and childhood.
Septic arthritis affects weight-bearing joints in three fourths of cases. When the disease occurs in infancy, joint dysfunction may not be apparent until many months later. We located 49 children who had had 50 episodes of septic arthritis from 1 1/2 to 12 years earlier (mean, 4.3 years). Thirteen patients (27%) had sequelae, and in eight (16%), there was impairment of ambulation. Residual damage was more common with hip and ankle involvement than with knee joint disease. Sequelae were equally common after Haemophilus influenzae and Staphylococcus aureus infection. Evaluation at the time of hosiptal discharge correctly identified only four of the 13 children with sequelae, and four others who were normal at follow-up had been thought to have permanent damage at discharge. Children with sequelae tended to have been sick longer before diagnosis, and drainage of pus was delayed.
Age Factors↗