Biomedical subjects

John D Martin

Publications and source records attributed to John D Martin.

6 recordsLinked to original sources

PubMed2006

The kinetic model of the shikimate pathway as a tool to optimize enzyme assays for high-throughput screening.

Four-enzyme section of the shikimate pathway (Aro B, D, E, and K) of Streptococcus pneumoniae has been studied. Kinetic properties of the individual enzymes and three- and four-enzyme linked reactions have been characterized in vitro. On the basis of the data measured in spectrophotometric and LC-MS experiments, kinetic mechanisms of the enzymes have been suggested and all kinetic parameters have been identified. Kinetic models for these three- and four-enzyme sections of the shikimate pathway have been constructed and validated. The model of the four-enzyme section of shikimate pathway has been employed to design an inhibition-sensitive reconstituted pathway for a high-throughput screening effort on the shikimate pathway. It was demonstrated that using the model it was possible to optimize this reconstituted pathway in such a way to provide equal sensitivity of the enzymes to inhibition.

Alcohol Oxidoreductases↗

PubMed2006

A second p53 binding site in the central domain of Mdm2 is essential for p53 ubiquitination.

Mdm2 negatively regulates p53 by inhibiting its transcriptional activity and promoting its degradation by functioning as an E3 ubiquitin ligase. The primary p53 binding site on mdm2 is located in its N-terminal domain. Through binding to p53 at its N-terminal transactivation domain, mdm2 directly blocks the transcriptional activation function of p53. We discovered that truncated mdm2 protein constructs without the N-terminal p53 binding domain are at least as active as full-length mdm2 in catalyzing p53 ubiquitination. Furthermore, the deletion of the central acidic domain significantly reduces the E3 ligase activity of mdm2 toward p53. We have also performed GST pull-down experiments to probe the direct binding of various mdm2 domain constructs toward full length p53 and found that mdm2 constructs without the N-terminal p53 binding domain retain the ability to bind to p53. Our kinetic and binding data localize the second p53 binding site between amino acids 211 and 361, including the acidic domain and the zinc finger region. Our work, consistent with other reports, suggests that the p53 tetramer interacts with at least two sites on mdm2. Although the interaction between the N-termini of mdm2 and p53 blocks the transactivation activity of p53, the interaction between the central domain of mdm2 and the core domain of p53 is critical for the ubiquitination and degradation of p53. This second mdm2-p53 interaction site represents an alternative target for small molecule modulators of the mdm2-p53 pathway.

Animals↗

PubMed2006

Remote endarterectomy: lessons learned after more than 100 cases.

BACKGROUND: Bypass procedures have been the mainstay of treatment of extensive external iliac and superficial femoral artery (SFA) occlusive disease, particularly total occlusions. Since the early 1990s, reports from Europe have espoused the virtues of endarterectomy of the superficial femoral and iliac arteries from a small groin incision, but adoption in the United States has been limited. Over the past 4 years, we have explored the technical challenges and durability of this procedure and report our findings. METHODS: Remote endarterectomy from an inguinal incision was the primary treatment option for all patients considered surgical candidates for vascular reconstruction of the external iliac and superficial femoral arteries. All data were entered into an outcomes database prospectively and reviewed retrospectively. After the procedure, duplex ultrasound surveillance was performed quarterly the first year and semi-annually thereafter. RESULTS: Remote endarterectomy was the planned procedure in 133 patients. The mean age was 68 years, 68% were men, and 31% were diabetic. The indications for the procedure were claudication in 57% and limb salvage in 43%. In 16 patients (12%), technical issues precluded the completion of the remote endarterectomy and a bypass was performed. Successful retrograde iliac endarterectomy was performed in 7 patients, SFA endarterectomy in 105 patients, and combined retrograde iliac and antegrade SFA in 5 patients. The average duration of the procedure was 162 minutes +/- 69 minutes (SD). Half of the patients were discharged on the first postoperative day, and the average length of stay was 2.52 days. The mean follow-up was 19 months, with a primary patency of 70% at 30 months by life-table analysis. Limb salvage was 94%. CONCLUSIONS: Remote endarterectomy is a viable and durable alternative to standard bypass procedures. It has equivalent patency to published results of bypass or endovascular procedures of the external iliac and superficial femoral arteries and may soon replace bypass as the preferred procedure for long-segment occlusions of these vessels.

Adult↗

PubMed2004

Remote superficial femoral artery endarterectomy and distal aSpire stenting: multicenter medium-term results.

OBJECTIVE: The purpose of this study was to examine the results of remote superficial femoral artery endarterectomy (RSFAE) in conjunction with distal aSpire stenting. METHODS: RSFAE is a minimally invasive procedure performed through a limited groin incision. Forty patients were included in the study. The indications for the procedure were claudication in 36 patients and limb salvage in 4 patients. RSFAE was performed with the MollRing Cutter device through a femoral arteriotomy. The distal atheromatous plaque was "tacked" with the aSpire stent, which is an expandable polytetrafluoroethylene-covered nitinol stent with high radial strength, yet is flexible and able to withstand compressive forces proximal to the knee joint. Before stent deployment, if the stent position is not optimal it can be wrapped down, repositioned, and re-expanded. Therefore, not only is the plaque end point tacked, but the collateral vessels may be preserved. All patients underwent follow-up examination with serial color-flow ultrasound scanning. RESULTS: The mean length of endarterectomized superficial femoral artery was 26.2 cm +/- 6.2 cm (range, 13-41 cm). The primary cumulative patency rate by means of life table analysis was 68.6% +/- 13.5% (SE) at 18 months (mean, 13.2 months; range, 1-31 months). During follow-up percutaneous transluminal balloon or stent angioplasty was necessary in 6 patients, for a primary assisted patency rate of 88.5% +/- 8.5% at 18 months. The locations of recurrent stenoses after RSFAE were evenly distributed along the endarterectomized artery. There were no deaths and one wound complication, and mean hospital length of stay was only 2.1 +/- 0.5 days. CONCLUSIONS: RSFAE with distal aSpire stenting is a safe and moderately durable procedure. If long-term patency rates are similar to those of above-knee femoropopliteal bypass graft, this procedure may prove to be a minimally invasive adjunct for the treatment of superficial femoral artery occlusive disease.

Adult↗

PubMed2003

A glass half full.

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Gelatin Sponge, Absorbable↗

PubMed2003

Production and characterization of monoclonal antibodies against the vasoconstrictive peptide human urotensin-II.

We report the production and characterization of four monoclonal antibodies (MAbs) against human urotensin-II (hU-II). The antibodies were raised against human hU-II, which contains the C-terminus cyclic ring (CFWKYC) that is conserved across species. Multiple selection assays were applied to ensure antibody potency and reactivity against the ring structure. The MAbs reacted via ELISA with hU-II bound to plastic, immunoprecipitated [(125)I-Y(9)] hU-II, bound to biotinylated hU-II in BIAcore analysis and, by Western analysis, recognized the full-length human preprourotensin-II expressed in transfected HEK293 cells. All four MAbs cross-reacted with porcine A, porcine B, rat, mouse, and goby U-II in ELISA. By competitive RIA, hU-II(5-11) (identical to the C-terminus of goby U-II) reacted equivalently to hU-II and goby U-II. The IC(50)s were 0.8 nM for one MAb and 1.6 nM for the others. All four MAbs reacted 15-fold less potently with hU-II(5-10) and 50-fold less potently with hU-II(5-10) amide. Thus, the ring structure and terminal Val/Ile comprise the binding site for this group of MAbs. This panel of antibodies could be useful tools to help delineate the biology and pharmacology of U-II. They may also be of diagnostic value in monitoring hU-II in body fluids.

Animals↗