PubMed Health⌕ Search

Biomedical subjects

B R Lee

Publications and source records attributed to B R Lee.

At least 19 recordsLinked to original sources
PubMed1995

Molecular cloning and sequence analysis of the scpZ gene encoding the serine carboxypeptidase of Absidia zychae.

Carboxypeptidase Z is a serine carboxypeptidase secreted by Absidia zychae NRIC 1199. The cDNA and genomic DNA carrying the scpZ gene encoding carboxypeptidase Z were cloned and sequenced. The nucleotide sequences of the cDNA (1.4 kb) and the genomic DNA (3.3 kb) were analyzed and the intervening sequences were located by a comparison of the two. It was found that the scpZ gene was interrupted by 11 short introns, 50-75 nucleotides in length. Genomic Southern analysis showed that there was only one scpZ gene in the genome of A. zychae. The gene encoded a putative pre-pro-enzyme composed of 409 amino-acid residues of the mature carboxypeptidase Z (M(r) 45,421) and an additional N-terminal sequence of 51 amino-acid residues. The amino-acid sequence around the active serine residue of carboxypeptidase Z (-G-E-S-Y-G-G-) differed from the consensus (-G-E-S-Y-A-G-) which is conserved in most of the serine carboxypeptidases so far analyzed.

Amino Acid Sequence↗
PubMed1995

Relationship between changes in prostate-specific antigen and the percent of prostatic epithelium in men with benign prostatic hyperplasia.

OBJECTIVES: Pretreatment knowledge of prostate gland histology would allow a more scientifically based selection of medical therapy for men with benign prostatic hyperplasia (BPH) and may increase the effectiveness of the pharmacologic agents available. Changes in prostate-specific antigen (PSA), or PSA velocity, may reflect prostatic epithelial growth in BPH. Our objective was to determine if PSA velocity prior to diagnosis correlated with the relative amount of epithelium in BPH tissue. METHODS: We evaluated 39 men with BPH who had serial PSA determinations (mean, 5.4) on frozen sera from 2.3 to 25.1 years before diagnosis, and archival material from simple prostatectomy available for pathologic evaluation. We used an immunoenzymatic staining technique for PSA to bind prostatic epithelium selectively so that color differences in the stained tissue sections could be used to quantify stroma, epithelium, and glandular lumina. RESULTS: The average percentage of epithelium (%E) was 12.4 and the average stroma-epithelial ratio (SER) was 6.6. The correlation of PSA velocity for the three visits nearest to prostatectomy (n = 32) versus %E and SER was significant (P = 0.003 for both). The PSA value nearest to prostatectomy (n = 39) was directly correlated with %E and SER (P = 0.0001 and P = 0.001, respectively). CONCLUSIONS: These data suggest that PSA and PSA velocity are directly related to the histologic makeup of the prostate in men with BPH. Thus, pretreatment evaluation of PSA could be useful as part of an evaluation to direct BPH therapy.

Aged↗
PubMed1995

A quantitative histological evaluation of the dilated ureter of childhood. II: Ectopia, posterior urethral valves and the prune belly syndrome.

A quantitative histological study was performed on specimens of 33 ureters obtained from 14 male and 19 female patients 5 days to 14 years old (mean age 1.2 years). A high resolution color image video analysis system was used to quantify and compare collagen and smooth muscle components of the muscularis layers to normal control ureters from patients of similar ages. In comparing ureters with ectopia (7), ureters with ectopic ureteroceles (20) and control ureters (4) there was not a statistically different collagen-to-smooth muscle ratio among the groups. In the patients with posterior urethral valves the amount of collagen and smooth muscle was not statistically different from controls (p > 0.01), although the collagen-to-smooth muscle ratio was increased. The 4 patients with the prune belly syndrome had a collagen-to-smooth muscle ratio that was markedly elevated (1.21 versus 0.39) compared with controls. When this group was analyzed as 2 separate groups (obstructed versus refluxing ureters) the difference was more apparent (p < 0.004). Ureters with ectopia or ectopic ureteroceles and ureters associated with posterior urethral valves had similar quantitative amounts of smooth muscle (60%, 56% and 52%, respectively). In patients with the prune belly syndrome obstructed ureters had 65% muscle and refluxing ureters had 38% muscle on evaluation. The percentage of collagen was 33% in ureters with ectopia, 37% in those with ureteroceles and 48% in those associated with posterior urethral valves compared with 23% in controls. In the group with the prune belly syndrome there was 30% collagen in obstructed ureters and 62% collagen in refluxing ureters. Our findings demonstrate that while these dilated ureters had different etiologies the overall quantitative composition of collagen-to-smooth muscle ratios was similar except in refluxing ureters associated with the prune belly syndrome. Our study provides further insight into the pathological nature of such ureters and considerations for surgical repair.

Adolescent↗
PubMed1994

Identification of the N-terminal domain of enzyme I of the Escherichia coli phosphoenolpyruvate:sugar phosphotransferase system produced by proteolytic digestion.

The phosphoenolpyruvate:sugar phosphotransferase system of bacteria plays an important role in the concomitant uptake and phosphorylation of numerous sugars. The first protein in the pathway of phosphotransfer of the phosphoenolpyruvate:sugar phosphotransferase system is Enzyme I. It has been shown that a stable N-terminal domain can be produced by treatment of the purified protein with various proteolytic enzymes. We show here that the region from glutamate-252 to leucine-264 is accessible to proteolysis resulting in N-terminal cores ranging from M(r) 27521 to 28799.

Amino Acid Sequence↗
PubMed1994

Radical prostatectomy for high grade disease: a reevaluation 1994.

Traditionally, patients with high grade tumors (Gleason score 8 to 10) were not considered candidates for radical prostatectomy because of poor long-term survival rate. However, with improvements in the staging of prostate cancer and a reduction in the morbidity of radical prostatectomy, it is reasonable to reevaluate the results of radical prostatectomy in high grade disease in a contemporary setting. We studied the clinical outcome of 72 men with Gleason scores 8 to 10 on needle biopsy who presented with clinically localized disease (T1c [9], T2a [22], T2b [17], T2c [13] and T3a [11]). Nine patients (13%) did not undergo radical prostatectomy because of positive lymph nodes identified on frozen section. Of the 63 men who underwent radical prostatectomy 43 (68%) had negative lymph nodes and 20 (32%) had positive lymph nodes. The actuarial likelihood of having an undetectable serum prostate specific antigen at 5 years was 43% for men with negative lymph nodes and 45% for men with specimen confined disease. In 7 men (9%) distant metastases developed and all had positive lymph nodes at surgery. These data suggest that men with high grade disease who are suitable candidates for radical prostatectomy should have the pelvic lymph nodes evaluated. If the lymph nodes are negative, the patient may benefit from an attempt at surgical cure.

Actuarial Analysis↗
PubMed1994

Concordance rates for benign prostatic disease among twins suggest hereditary influence.

OBJECTIVES: The etiology of benign prostatic hyperplasia (BPH) is unknown. Evidence for a hereditary trait would provide new avenues for investigation. METHODS: We compared the concordance for benign prostate disease in monozygotic (MZ) and dizygotic (DZ) twins who served in the United States military in World War II and have been followed by the Medical Follow-up Agency, Institute of Medicine of the National Academy of Sciences. Questionnaires completed in 1985 by 10,000 twins were reviewed for evidence of prostatic disease. Key words indicating benign prostatic disease (prostate, prostatectomy, BPH, TURP, and prostatism) were identified in 533 (5.3%) of the questionnaires. RESULTS: The average age was 64 +/- 3 years (range 56 to 68 in 1985). After eliminating men with known prostate cancer, there were 256 twin pairs that were informative for benign prostatic disease: both twins were concordant in 25 instances and discordant in 231, with only one twin mentioning benign prostatic disease. The pairwise concordance for MZ twins was 14.7% (19 of 129) and for DZ pairs it was 4.5% (5 of 112). The relative risk for benign prostatic disease for MZ twins was thus 3.3 (p = 0.008). The probandwise concordance rates, which express the probability of BPH in a cotwin of an affected twin, were 25.7% for MZ twins and only 8.5% for DZ twins. A covariance analysis estimated that 49% of the observed variance between twins could be attributed to genetic effects. CONCLUSIONS: These data provide preliminary evidence for the heritability of benign prostatic disease.

Aged↗
PubMed1993

Purification and characterization of serine carboxypeptidases from Absidia zychae.

Among 29 strains of zygomycetes screened for serine carboxypeptidases, Absidia zychae NRIC 1199 showed the highest enzyme production. Two serine carboxypeptidases, CPZ-1 and CPZ-2, were purified to a homogeneous state from an extract of koji culture of A. zychae NRIC 1199. Purified CPZ-1 and CPZ-2 showed similar properties except the isoelectric point (pI); The pI of CPZ-1 and CPZ-2 were 4.50 and 4.65, respectively. The molecular weights of the CPZ-1 and CPZ-2 were 48,000 by SDS-PAGE and gel filtration. Among the proteinase inhibitors tested, phenylmethylsulfonyl fluoride and monoiodoacetic acid strongly inhibited the enzyme activity. The optimum pHs of CPZ-1 and CPZ-2 were 4.2 towards Z-Glu-Tyr. It is shown that the substrate specificities of CPZ-1 and CPZ-2 were dependent on the presence of bulky amino acid residues in the penultimate position (P1) for the small Z-peptides. However, in spite of the presence of Gly, Asp, Arg, or Pro in the P1 position, oligopeptides were hydrolyzed rapidly. CPZ-1 and CPZ-2 had not only carboxypeptidase but also carboxyamidase and amidase activities, and acted preferentially as a carboxyamidase for C-terminal amidated peptides. The hydrophobicity of P2 and P3 positions and the bulkiness of P1 and P'1 positions of the substrate may be important for carboxyamidase and amidase activities.

Amino Acid Sequence↗
PubMed1992

Plasma lipid transport in the preruminant calf, Bos spp: primary structure of bovine apolipoprotein A-I.

The preruminant calf (Bos spp.) is a model of considerable interest with regard to hepatic and intestinal lipoprotein metabolism (Bauchart et al., J. Lipid Res. (1989) 30, 1499-1514 and Laplaud et al., J. Lipid Res. (1990) 31, 1781-1792). As a preliminary step towards future experiments dealing with HDL metabolism in the calf, we have purified apoA-I from this animal and determined its complete amino acid sequence. Thus, approx. 10% of calf apoA-I was shown to contain a propeptide, with the sequence Arg-His-Phe-Trp-Gln-Gln. Enzymatic cleavage of apoA-I resulted in 10 proteolytic peptides. The complete apoA-I sequence was obtained after alignment of peptides on the basis of their homologies with those from rabbit apoA-I. Thus calf apoA-I consists of 241 amino acid residues, and exhibits high sequence homology with all mammalian apoA-I's studied to date. The bovine protein contained 10 hydrophobic amphipathic helical regions, occurring between residues 43-64, 65-86, 87-97, 98-119, 120-141, 142-163, 164-184, 185-206, 207-217 and 218-241. A computer-constructed phylogenetic tree showed that bovine apoA-I was more closely related to its dog counterpart, including the presence of a single methionine, than to the corresponding macaque and human proteins. Comparative predictions of the respective antigenic structures of human and bovine apoA-I's using the Hopp-Woods algorithm indicated similar positions for all 13 detectable antigenic sites, among which 7 were of identical, or closely related, amino acid composition. This finding was confirmed by demonstration of partial immunological identity between the two proteins upon immunodiffusion analysis, a result obtained using a monospecific rabbit antiserum against bovine apoA-I. Finally, comparison of sequence homology between bovine apoA-I and the lecithin:cholesterol acyl transferase (LCAT) activating region of human apoC-I suggests that several LCAT activating domains may be present in calf apoA-I.

Amino Acid Sequence↗
PubMed1992

A quantitative histological analysis of the dilated ureter of childhood.

A quantitative histological study of the dilated ureter of childhood was performed on 26 ureters. The specimens were from 15 male and 11 female patients 10 days to 12 years old (mean age 2.0 years). A color image analysis system was used to examine and compare collagen and smooth muscle components of the muscularis layers to normal control ureters of similar age. In comparing primary obstructed (12) to primary refluxing (14) megaureters and control ureters (6), there was a statistically different collagen-to-smooth muscle ratio (p < 0.001) between the primary obstructed and primary refluxing megaureter groups. For patients with primary refluxing megaureter there was a 2-fold increase in the tissue matrix ratio of collagen-to-smooth muscle when compared to patients with primary obstructed megaureter. In the primary obstructed megaureters the amount of collagen and smooth muscle was not statistically different from controls (p > 0.01). The increased tissue matrix ratio of 2.0 +/- 0.35 (collagen-to-smooth muscle) in the refluxing megaureter group compared to 0.78 +/- 0.22 in the obstructed megaureter group and 0.52 +/- 0.12 in controls was found to be due not only to a marked increase in collagen but also a significant decrease in the smooth muscle component of the tissue. Primary obstructed and normal control ureters had similar quantitative amounts of smooth muscle with 60 +/- 5% and 61 +/- 6%, respectively, while refluxing megaureters had only 40 +/- 5% smooth muscle. The percentage collagen was 36 +/- 5 in the obstructed megaureter group and 30 +/- 5 in controls, with refluxing megaureters having 58 +/- 5% collagen on analysis. Our findings emphasize the significant differences in the structural components (collagen and smooth muscle) of the dilated ureter of childhood, and provide us with further insight into the pathological nature of these dilated ureters and their surgical repair.

Child↗
PubMed1991

The leader sequence of streptokinase is responsible for its post-translational carboxyl-terminal cleavage.

When the expression of streptokinase from two tac promoter-controlled expression vectors, one of these deleted a putative leader sequence of streptokinase and the other not, was compared, both normal and degraded streptokinase were detected in proteins expressed from the leader-contained vector, but only normal streptokinase was detected from the leader-deleted vector. These findings indicate that the characteristic carboxyl-terminal cleavage of streptokinase is correlated with its leader sequence and occurs during the defective secretion. The homogeneous preparation of streptokinase was facilitated by expressing from this leader-deleted vector.

Base Sequence↗
PubMed1991

Amino acid sequence of rabbit apolipoprotein E.

The complete amino acid sequence of rabbit apolipoprotein E (apoE) was determined by generating three sets of peptides using cyanogen bromide, endoproteinase AspN, and Staphylococcus aureus V8 protease to cleave the protein. Through twenty cycles of sequence analysis on the whole protein, glutamic acid was identified as the N-terminal residue of rabbit apoE; the C-terminus of the protein was identified as glutamine. Based on the sequence of 294 amino acid residues determined by protein structure analysis, the molecular weight of rabbit apoE was determined to be 33,684. The protein sequence differed from the cDNA inferred sequence in 19 positions, only one of which could be attributed to microheterogeneity. The corrected amino acid sequence of rabbit apoE shares 80% homology with the human apoE sequence, 4% greater homology than that inferred from the cDNA sequence. The great similarity in the amino acid sequences of human and rabbit apoE suggests that their physical and physiological properties may also be similar. This homology and the relative ease with which apoE is isolated from rabbit plasma make it possible to conduct some in vitro experiments with the rabbit apoprotein that would have direct relevance to human apoE, but would be difficult or impossible with the human counterpart because of the quantity of protein required.

Amino Acid Sequence↗
PubMed1990

Isolation and characterization of sulfhydryl and disulfide peptides of human apolipoprotein B-100.

Twenty-three of the 25 cysteine residues in apolipoprotein B-100 have been isolated directly from tryptic or peptic peptide mixtures. Sixteen cysteine residues exist in disulfide forms: Cys-1-Cys-3, Cys-2-Cys-4, Cys-5-Cys-6, Cys-7-Cys-8, Cys-9-Cys-10, Cys-11-Cys-12, Cys-13-Cys-14, and Cys-20-Cys-21. All of these except Cys-20-Cys-21 are recently discovered disulfide linkages. In addition to Cys-22 and Cys-24, which have been described as sulfhydryls on low density lipoprotein, Cys-15 to Cys-18 and Cys-23 are in the reduced form. Cys-19 and Cys-25 are not yet confirmed. Our results revealed that all identified disulfide linkages are located in the trypsin-releasable regions and that all except Cys-1-Cys-3 and Cys-2-Cys-4 are linked to the neighboring cysteine. We propose a linear model of apolipoprotein B-100 in low density lipoprotein that wraps around the low density lipoprotein molecule.

Amino Acid Sequence↗
PubMed1989

Site-specific alteration of Gly-24 in streptokinase: its effect on plasminogen activation.

Oligonucleotide-directed mutagenesis was carried out to replace glycine-24 of streptokinase with histidine, glutamic acid, or alanine. Substitutions with either histidine or glutamic acid resulted in almost complete loss of streptokinase activity but streptokinase replaced with alanine retained its activity. Although streptokinases with histidine-24 or glutamic acid-24 bound normally to human plasminogen, they were not able to generate active plasmin, whereas those with alanine-24 or glycine-24 (wild-type) could generate active plasmin. The results indicate that the small, uncharged alkyl group side-chain on the 24th amino acid residue of streptokinase is indispensable for the activity of the human plasminogen-streptokinase complex.

Alanine↗
PubMed1989

The primary structure of human apolipoprotein A-IV.

Human apolipoprotein (apo) A-IV was purified from chylous ascites fluid. Proteolytic peptides produced by trypsin and Staphylococcus aureus V8 proteinase digestions were purified by high-performance liquid chromatography and sequenced. Human apoA-IV contains 376 amino acid residues. The peptide-derived sequence generally matches two previously reported DNA-derived amino acid sequences except for discrepancies in five positions. In order to examine these discrepancies further, one complete apoA-IV cDNA clone and another partial clone were sequenced. Comparison of all the available information indicates that the peptide-derived sequence reported here is accurate. Sequencing errors probably account for some of the discrepancies between the two primary sequences predicted by earlier nucleotide analyses. In certain positions, however, bona fide sequence heterogeneity or cloning artifact cannot be excluded.

Amino Acid Sequence↗
PubMed1989

A digital image microscopy system for rare-event detection using fluorescent probes.

Instrumentation for rare-event analysis should be capable of reliably detecting infrequent cells (less than 1:10,000) while both excluding false-positive signals and including true positive cells found in multicell clumps. We have developed a digital image microscopy (DIM) system in which a cytospin of 2 million cells is scanned with an intensified video camera (ISIT) using an IBM PC AT microcomputer-controlled microscope stage. PASCAL software controls the stage and analyzes video input, storing the location of positive cells to magnetic disk. The user can then "replay" each positive cell under computer control for either visual confirmation or analysis using other fluorescent probes. The computer requires 24 min to scan a cytoprep of 2 million cells, while playback for visual confirmation by the user averages 5 min. Using Hoechst-33342 premarked cells seeded into bone marrow as a model system, we found that the DIM system reliably detects one target cell per million marrow cells. With appropriate immunological markers, this system will aid in evaluating bone marrow purged of tumor cells prior to transplantation and should also be useful for detection of minimal residual disease in blood or bone marrow from patients with leukemia or solid tumors.

Bone Marrow↗
PubMed1988

Microcomputer software and interface for control of a microscope scanning stage.

Motorized scanning stages are valuable in microscopy systems that employ digital image analysis and for the development of semiautomatic computer-assisted microscope systems; the development of standard software "tools" to control such stages will facilitate their integration into a variety of computer-based systems. A set of Microsoft BASIC and Turbo PASCAL programs that interfaces a microprocessor-controlled stepper motor microscope stage (MDACE 1000) to an IBM PC or PC-AT or compatible microcomputer via a serial interface (RS-232) is described. These programs can be integrated into other software written in either BASIC or PASCAL, or used via a menu program that directs the routines to control scanning patterns and to locate the microscope stage to a selected area of the slide. Coordinates of significant events on a slide can be stored on a disk file to allow future examination. The software and interface also provide control of a filter wheel in the microscope for use in multicolor fluorescence assays.

Computers↗