Structure and function of enzymes involved in the biosynthesis of phenylpropanoids.

As a major component of plant specialized metabolism, phenylpropanoid biosynthetic pathways provide anthocyanins for pigmentation, flavonoids such as flavones for protection against UV photodamage, various flavonoid and isoflavonoid inducers of Rhizobium nodulation genes, polymeric lignin for structural support and assorted antimicrobial phytoalexins. As constituents of plant-rich diets and an assortment of herbal medicinal agents, the phenylpropanoids exhibit measurable cancer chemopreventive, antimitotic, estrogenic, antimalarial, antioxidant and antiasthmatic activities. The health benefits of consuming red wine, which contains significant amounts of 3,4',5-trihydroxystilbene (resveratrol) and other phenylpropanoids, highlight the increasing awareness in the medical community and the public at large as to the potential dietary importance of these plant derived compounds. As recently as a decade ago, little was known about the three-dimensional structure of the enzymes involved in these highly branched biosynthetic pathways. Ten years ago, we initiated X-ray crystallographic analyses of key enzymes of this pathway, complemented by biochemical and enzyme engineering studies. We first investigated chalcone synthase (CHS), the entry point of the flavonoid pathway, and its close relative stilbene synthase (STS). Work soon followed on the O-methyl transferases (OMTs) involved in modifications of chalcone, isoflavonoids and metabolic precursors of lignin. More recently, our groups and others have extended the range of phenylpropanoid pathway structural investigations to include the upstream enzymes responsible for the initial recruitment of phenylalanine and tyrosine, as well as a number of reductases, acyltransferases and ancillary tailoring enzymes of phenylpropanoid-derived metabolites. These structure-function studies collectively provide a comprehensive view of an important aspect of phenylpropanoid metabolism. More specifically, these atomic resolution insights into the architecture and mechanistic underpinnings of phenylpropanoid metabolizing enzymes contribute to our understanding of the emergence and on-going evolution of specialized phenylpropanoid products, and underscore the molecular basis of metabolic biodiversity at the chemical level. Finally, the detailed knowledge of the structure, function and evolution of these enzymes of specialized metabolism provide a set of experimental templates for the enzyme and metabolic engineering of production platforms for diverse novel compounds with desirable dietary and medicinal properties.

Structure and mechanism of chalcone synthase-like polyketide synthases.

Polyketide synthases (PKS) produce an array of natural products with different biological activities and pharmacological properties by varying the starter and extender molecules that form the final polyketide. Recent studies of the simplest PKS, the chalcone synthase (CHS)-like enzymes involved in the biosynthesis of flavonoids, anthocyanin pigments, and antimicrobial phytoalexins, have yielded insight on the molecular basis of this biosynthetic versatility. Understanding the structure-function relationship in these PKS provides a foundation for manipulating polyketide formation and suggests strategies for further increasing the scope of polyketide biosynthetic diversity.

Structural control of polyketide formation in plant-specific polyketide synthases.

BACKGROUND: Polyketide synthases (PKSs) generate molecular diversity by utilizing different starter molecules and by controlling the final length of the polyketide. Although exploitation of this mechanistic variability has produced novel polyketides, the structural foundation of this versatility is unclear. Plant-specific PKSs are essential for the biosynthesis of anti-microbial phytoalexins, anthocyanin floral pigments, and inducers of Rhizobium nodulation genes. 2-Pyrone synthase (2-PS) and chalcone synthase (CHS) are plant-specific PKSs that share 74% amino acid sequence identity. 2-PS forms the triketide methylpyrone from an acetyl-CoA starter molecule and two malonyl-CoAs. CHS uses a p-coumaroyl-CoA starter molecule and three malonyl-CoAs to produce the tetraketide chalcone. Our goal was to elucidate the molecular basis of starter molecule selectivity and control of polyketide length in this class of PKS. RESULTS: The 2.05 A resolution crystal structure of 2-PS complexed with the reaction intermediate acetoacetyl-CoA was determined by molecular replacement. 2-PS and CHS share a common three-dimensional fold, a set of conserved catalytic residues, and similar CoA binding sites. However, the active site cavity of 2-PS is smaller than the cavity in CHS. Of the 28 residues lining the 2-PS initiation/elongation cavity, four positions vary in CHS. Point mutations at three of these positions in CHS (T197L, G256L, and S338I) altered product formation. Combining these mutations in a CHS triple mutant (T197L/G256L/S338I) yielded an enzyme that was functionally identical to 2-PS. CONCLUSIONS: Structural and functional characterization of 2-PS together with generation of a CHS mutant with an initiation/elongation cavity analogous to 2-PS demonstrates that cavity volume influences the choice of starter molecule and controls the final length of the polyketide. These results provide a structural basis for control of polyketide length in other PKSs, and suggest strategies for further increasing the scope of polyketide biosynthetic diversity.

Bilateral sternocleidomastoid tumors of infancy.

The sternocleidomastoid tumor of infancy (STOI) is a relatively uncommon condition. Typically, it presents as a firm, well circumscribed mass within the sternocleidomastoid muscle (SCM) in infants 1-8 weeks of age and may be associated with torticollis. This condition must be considered in any infant with a lateral neck mass. The diagnosis can often be made clinically, but unusual presentations may present diagnostic challenges. Although bilateral involvement is rare, it does occur. The second reported case, a 2-week old female with bilateral STOIs and torticollis, is reported. Although many of the characteristics of the masses suggested the condition, the bilateral nature added uncertainty to the clinical impression, and magnetic resonance imaging (MRI) was used to confirm the diagnosis. The clinical presentation and management of the STOI are reviewed, and the unusual features of this case are discussed.

Complications of frontal sinusitis in adolescents: case presentations and treatment options.

Frontal sinusitis in adolescents can be a serious condition because of the severity of potential complications and rapid progression of disease. These complications may arise as a result of extension of the infection intracranially, to the frontal bone, and to the orbit. Infection may spread hematogenously or by direct extension. A series of seven adolescent patients with complications of frontal sinusitis is presented. Surgical options are necessitated by the location and extent of infection and include trephination, frontal sinus obliteration, and craniotomy. Recent advances in endoscopic sinus surgery have provided surgeons another access to the frontal recess. However, a combined otolaryngological/neurosurgical approach is essential for patients with intracranial complications.

Management of the carotid artery following penetrating injuries of the soft palate.

Penetrating injuries of the soft palate are not uncommon in the pediatric population. The majority are minor, requiring only conservative treatment. Despite the close proximity of the carotid artery to lateral soft palate and tonsillar fossa, the incidence of carotid injury is low. When carotid injury does occur, it is usually secondary to blunt trauma to the carotid with a resultant thrombosis. A concomitant neurologic deterioration often follows occurring from 3 h to 3 days after the initial injury (Hengerer et al. (1984). Laryngoscope 94, 1571-1575). The potential neurologic sequelae of such an injury make these seemingly innocuous wounds a diagnostic and management dilemma for the clinician. We present an unusual case of an internal carotid artery pseudoaneurysm in a neurologically intact child following soft palate impalement. We review the diagnostic evaluation including angiography and magnetic resonance angiography, the surgical approach, and postoperative issues including anticoagulation. The literature is reviewed and a discussion regarding the approach to penetrating injuries of the soft palate is presented.

Pediatric cervical esophageal perforation secondary to abusive blunt thoracic trauma.

Cervical esophageal perforation secondary to blunt trauma is extremely rare. A case of an 8-week-old infant that sustained a cervical esophageal perforation from abusive acceleration/deceleration blunt trauma is presented. Early diagnosis and appropriate intervention of an already insidious injury can be further delayed by the lack of concise temporal events and falsification of data from the abusive caretaker(s). In-depth history taking, documentation and notification of authorities lays the critical medicolegal groundwork for this unfortunately increasing problem in our society. Diagnosis and management are discussed. The pediatrician, emergency department, nursing staff and otolaryngologist should have strong suspicions of abuse when esophageal perforation is identified in an infant.

Cunninghamella bertholletiae and Pneumocystis carinii pneumonia as a fatal complication of chronic lymphocytic leukemia.

A unique patient with chronic lymphocytic leukemia died with pneumonia caused by both Cunninghamella bertholletiae and Pneumocystis carinii. In tissue sections, the hyphae of C bertholletiae were twisted and ribbon-like, but were smaller than those typical for zygomycetes, displayed more than occasional septa, and exhibited Y branching, making histologic distinction from Aspergillus sp difficult.

Serous surface papillary carcinoma. A clinicopathologic study of 10 cases and comparison with stage III-IV ovarian serous carcinoma.

We compared 10 patients with serous surface papillary carcinoma (SSPC) to a control group of 16 patients with serous ovarian papillary carcinoma (SOPC). By definition, the ovaries were of normal size and shape in SSPC. In six cases, however, at least one ovary had focal surface irregularities. Microscopically, SSPC and SOPC differed only in the nature of their ovarian involvement. SSPC was confined to the ovarian surface or showed focal minimal invasion. Four patients with SSPC had coexistent, nonperitoneal serous-type carcinomas; three had superficial endometrial adenocarcinomas; and one had intramucosal adenocarcinoma of the fallopian tube. There were no associated carcinomas in the SOPC group. SSPC had a significantly worse prognosis than SOPC, as measured by median disease-free interval (SSPC: 3 months, SOPC: 9 months), median survival (SSPC: 12 months, SOPC: 24 months), and predicted 5-year survival (SSPC: all dead at 52 months, SOPC: 22.7%). SSPC is a clinicopathologically distinctive variant of serous carcinoma. Because of its more aggressive clinical course, it should be distinguished from conventional SOPC with peritoneal implants.

Impaired left ventricular function during exercise in coronary artery disease and exertional hypotension.

To characterize the hemodynamic abnormalities responsible for exertional hypotension coronary artery disease, we studied 11 patients with exertional hypotension during supine cycle ergometer exercise, defined as greater than 10 mm Hg decrease in systolic blood pressure during exercise, and 11 patients without exertional hypotension (controls). Patients were similar with respect to age, left ventricular ejection fraction at rest, and the intensity of exercise relative to maximal treadmill exercise capacity. Peak exercise ejection fraction, determined by radionuclide ventriculography, was significantly lower in patients with, than in those without exertional hypotension (50 +/- 3 vs. 56 +/- 3%; p less than 0.025). Ejection fraction and stroke volume decreased with exercise in patients with exertional hypotension but not in the controls even though changes in end-diastolic volume and mean blood pressure were similar in both groups. Peak exercise systolic blood pressure and rate pressure product were significantly lower in the patients with exertional hypotension than those without. The exercise-induced regional left ventricular contraction abnormalities were more prominent, extensive and frequent in patients with exertional hypotension than controls. Impairment of left ventricular contractile function was further evident by an abnormal end-systolic volume-systolic blood pressure relation in patients with exertional hypotension. These patients attained a much smaller increase in systolic blood pressure compared with controls despite no statistically significant differences in end-systolic volume response to exercise. These findings suggest that exertional hypotension in patients with ischemic heart disease is associated with exercise-induced left ventricular systolic dysfunction secondary to extensive myocardial ischemia.

Oncocytoid adenocarcinoma of the parotid gland. Cytologic, histologic and ultrastructural findings.

An oncocytoid adenocarcinoma of the parotid gland, diagnosed by fine needle aspiration cytology, is reported. The neoplasm, seen in a 66-year-old man, metastasized to 33 of 46 resected cervical lymph nodes; the patient is currently free of disease 17 months after surgery. Cytologically, the neoplastic cells occurred singly and in small clusters. They had abundant granular cytoplasm with occasional vacuoles, large nuclei and prominent nucleoli. Ultrastructurally, the tumor cells had copious cytoplasm, with moderately increased numbers of mitochondria, dilated endoplasmic reticulum, mucin droplets and intracytoplasmic lumina. The clinical importance of distinguishing this neoplasm from true malignant oncocytoma is, at present, unknown.

Angiomatoid carcinoma of the thyroid gland. Anaplastic carcinoma with follicular and medullary features mimicking angiosarcoma.

A 79-year-old man with a 28-year history of a substernal goiter had an anaplastic thyroid neoplasm with prominent angiomatoid features develop. Based on the light microscopic appearance of the tumor and the presence of immunocytochemical staining for Factor VIII-related antigen, a diagnosis of angiosarcoma was made. Subsequent immunocytochemical and ultrastructural studies suggested follicular and medullary differentiation without staining for Ulex europeaus I lectin or evidence of Weibel-Palade bodies. Based on these findings, the tumor is interpreted as an anaplastic carcinoma with evidence of mixed follicular and medullary differentiation and extensive intravascular growth producing an angiomatoid appearance. Positivity for Factor VII-related antigen may be due to nonspecific uptake of antigen-rich serum and platelets by the phagocytic tumor cells. Factor VIII-related antigen immunoreactivity should be interpreted with caution in intravascular neoplasms.

Pleural malignant mesothelioma following Wilms' tumor.

A 28-year-old woman had a left pleural malignant mesothelioma develop, which resulted in her death. At age four, she had undergone a left nephrectomy for Wilms' tumor and had received radiation therapy to the left renal fossa and to the right lung, the latter for a presumed diagnosis of metastatic tumor. Asbestos body counts of digested lung tissue were within the normal range. This is the fourth reported case of malignant mesothelioma following Wilms' tumor and is the first to provide quantitative analysis of asbestos in the lung.

Lymphoepithelioma-like carcinoma of the uterine cervix. A distinctive, undifferentiated carcinoma with inflammatory stroma.

Two cytologically uniform, light-microscopically undifferentiated carcinomas of the uterine cervix are described. The tumors were morphologically identical to nasopharyngeal lymphoepitheliomas, including the presence of an intense inflammatory stromal reaction with prominent lymphocytes, eosinophils, and plasma cells. One neoplasm occurred in a 29-year-old, was clinically Stage IB, and was successfully treated with radiation therapy, with 10-year disease-free follow-up. The second tumor developed in a 58-year-old, was clinically Stage IIIB, and resulted in the patient's death 17 months after diagnosis. When the malignant cells in these tumors were widely separated by inflammation, they could be easily overlooked or confused with lymphoproliferative lesions. Immunocytochemical stains were performed on one case. The tumor cells stained strongly for keratin and epithelial membrane antigen, but were negative for leukocyte common antigen, verifying their epithelial nature. Until the biologic behavior of this cytologically distinctive tumor is more clearly understood, it should be separated from conventional cervical cancers with prominent stromal inflammation.

The effect of left ventricular systolic function on maximal aerobic exercise capacity in asymptomatic patients with coronary artery disease.

The purpose of this study was to examine the relationship between maximal O2 uptake (VO2max) and left ventricular systolic function in patients with coronary artery disease. We studied 27 patients, age 50 +/- 10 years (mean +/- SD), who were asymptomatic and able to attain true VO2max. VO2max was defined by the leveling-off criterion and/or a respiratory exchange ratio of 1.15 or greater. Left ventricular ejection fraction was determined by gated cardiac blood pool imaging. In patients whose ejection fraction decreased with exercise, VO2max was 21 +/- 4 vs 27 +/- 4 ml/kg/min in those whose ejection fraction increased (p less than .001). Systolic blood pressure/end-systolic volume relation was shifted upward and to the right in the former group in response to peak exercise. In contrast, the pressure-volume relation was shifted upward and to the left in patients whose ejection fraction increased with exercise. Ejection fraction at rest did not correlate with VO2max. There was a significant but weak correlation between peak exercise ejection fraction and VO2max (r = .43, p less than .025). Left ventricular exercise reserve, i.e., the change in ejection fraction from rest to exercise, correlated with VO2max (r = .77, p less than .0002), maximal O2 pulse (r = .50, p less than .005), and maximal heart rate during treadmill exercise (r = .61, p less than .001). Maximal heart rate during treadmill exercise correlated with VO2max (r = .70, p less than .0002).(ABSTRACT TRUNCATED AT 250 WORDS)