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Biomedical subjects

C Grandi

Publications and source records attributed to C Grandi.

At least 37 records · Page 2Linked to original sources
PubMed1983

Spectroscopic study on the structure and stability of beef liver arginase.

The secondary and tertiary structure of the oligomeric arginase (EC 3.5.3.1) from beef liver was investigated by circular dichroism (CD) and fluorescence measurements. The far-ultraviolet CD spectrum of the enzyme at neutral pH is indicative of high helical content. The intrinsic fluorescence emission of the protein is due to tryptophan, the contribution of tyrosine being small. Upon excitation at 295 nm, the maximum of emission occurs at 330 nm, implying that the tryptophan residues are rather buried in a hydrophobic interior of the protein. Ethylenediaminetetraacetic acid (EDTA), which inactivates the enzyme by removing the functional Mn2+-ion from the enzyme, does not dissociate the enzyme into subunits, nor affect noticeably its secondary and tertiary structure. Inactivation occurs in the acid pH range, being complete at pH below 4. However, acidification up to pH 1.5 produced only limited changes in the far-ultra-violet CD spectrum and intrinsic fluorescence emission properties. The enzyme shows noteworthy thermal stability, as shown by measuring the residual activity after heating and by evaluating the temperature dependence of the CD signal at 220 nm and the intensity of emission fluorescence. A temperature of half inactivation (Tm) of 77 degrees was determined upon heating the enzyme at pH 7.5 in the presence of Mn2+-ions for 10 min; in the presence of EDTA, Tm is shifted to 55 degrees. Taken together, these observations indicate that the structural stability of beef liver arginase arises from a clustering of hydrophobic amino acids and from Mn2+-ion binding.

Animals↗
PubMed1982

Secondary lung cancer resection with curative intent: causes of success and failure and prognostic factors.

Observed and disease-free survivals were evaluated in a consecutive series of 46 resections of pulmonary metastases, with major chance of being a unique phenomenon. Survival curves were computed both since the treatment of primary tumors and since resection of lung metastases. From the treatment of primary tumors, median disease-free interval was 33 months, and rose to 66 months after resection of lung metastases. From the treatment of secondary lung cancers the observed survivals at 1, 3 and 5 years were respectively 60%, 41% and 26%. Survival was clearly affected by development and resectability of post-thoracotomic recurrence (100% without recurrence, 50% with resectable recurrence and 4% with unresectable recurrence). Recurrence rate was related to the first disease-free interval and to the anatomical extent (particularly for nodal status) of secondary lung cancer. This fact suggests that the failure of secondary lung cancer resection may arise either from the primary cancer (poor selection) or from the secondary cancer (delay in treatment).

Adolescent↗
PubMed1981

Micellar solubilization of biopolymers in organic solvents. Activity and conformation of lysozyme in isooctane reverse micelles.

Lysozyme solubilized in reverse micelles of bis(2-ethylhexyl) sodium sulfosuccinate in isooctane containing as little as 0.8% water (v/v) has been shown to be active. The amount of enzymatic activity depends on the water content and the pH of the reverse micellar system and under optimum conditions (i.e. pH 7.7 with 1.2% water) is 90% of the activity in water. The dependence of lysozyme activity on pH in reverse micelles is different than that in water, with the entire pH profile shifted 2 to 3 pH units higher in reverse micelles. Moreover, maximum enzyme activity is not found at the highest water contents tested (i.e. 1.6% and 2.0% water), but instead at 1.2% water. The Km for the N-acetylglucosamine oligomers used as substrate is 0.1 mM in reverse micelles (compared to 0.01 mM in water) when the concentration of substrate is referred to the water pools. Spectroscopic studies (CD, fluorescence, and UV absorbance) indicate that the conformation of lysozyme is significantly different in reverse micelles compared to water. In particular, CD studies indicate that the helical content of lysozyme changes from approximately 34% in water to approximately 48% in reverse micelles. Conformation and activity data are qualitatively correlated to the anomalous character of water in the reverse micelles. In particular, this may induce a stronger hydrogen bonding within the lysozyme which would in turn increase both the pKa of certain amino acid residues and the helical content of the macromolecule.

Animals↗
PubMed1980

Fluorescence properties of neutral protease from Bacillus subtilis.

The fluorescence properties of neutral protease from B. subtilis have been investigated under a variety of conditions and the results compared with those previously reported for the homologous metalloendopeptidase thermolysin (Fontana et al., 1977). In the pH range 5-9 neutral protease displayed a quite unusual fluorescence emission spectrum with a maximum near 320 nm, when excitation was at 295 nm. At this wavelength the protein fluorescence is due to tryptophan residues only, which, considering their blue-shift emission, appear rather buried in the hydrophobic protein interior. Specific removal of the functional zinc ion from the enzyme with tetraethylenepentamine does not lead to alteration of the microenvironment around tryptophan residues. On the other hand, removal of both zinc and calcium with ethylenediaminetetraacetic acid brings these residues in full contact with the aqueous solvent medium. Fluorescence quenching measurements were also used to determine the exposure of tryptophan residues in the native enzyme as well as in the presence of chelating agents and protein denaturants. Melting profile experiments carried out by monitoring the fluorescence intensity at 320 nm indicated a cooperative transition at 60-70 degrees. Temperature effects were also determined under conditions perturbed with respect to pH, guanidine hydrochloride and chelating agents. The results reveal differences in the fluorescence properties of the tryptophanyl residues of B. subtilis neutral protease relative to those of thermolysin, which are interpretable considering the location of these residues in the sequences of the two homologous proteins.

Amino Acid Sequence↗
PubMed1980

Preoperative irradiation and surgery for esophageal cancer: causes of failure.

Twenty-seven patients with squamous esophageal cancer underwent small volume, low-dose, concentrated radiotherapy followed by esophageal resection whenever possible (esophagectomy for tumors of the thoracic esophagus and esophagogastrectomy for tumors of the lower esophagus). Curative resectability was 70% (19/27) with 4 operative deaths (21%). Recurrence rate was 66% after a mean period of 16 months, and the failure pathway was nodal in 53% of the cases. Historical comparison of the data suggests that preoperative irradiation increases the curative resectability rate without changing the early recurrence rate or failure pathway. Tumors with deeper invasion of the esophageal wall, which benefit by preoperative irradiation, are probably related to greater nodal diffusion, which is partly outside of the volume that may be resected or irradiated.

Adult↗
PubMed1980

Thermolysin and Bacillus subtilis neutral protease. Conformation and stability of two homologous neutral metalloendopeptidases.

A comparative study on thermolysin from Bacillus thermoproteolyticus and neutral protease from Bacillus subtilis involving (far- and near-ultraviolet circular dichroism (CD) and immunological techniques is reported. These enzymes are homologous metalloendopeptidases, having similar size, kinetic behaviour, substrate specificity and susceptibility to inhibitors. The far-ultraviolet CD spectrum of each protein shows a minimum at 208 and a shoulder near 220 nm; differences in the extent of ellipticity, however, have been observed. Estimates of secondary structure obtained by quantitation of the far-ultraviolet CD spectra indicated a higher helicity of neutral protease relative to thermolysin. In the presence of ethylenediaminetetraacetic acid, which removes calcium and the functional zinc ion from the metalloenzymes, neutral protease is immediately denatured, whereas thermolysin maintains a globular structure, although thermolabile. On the other hand, the zinc-specific chelating agent tetraethylenepentamine does not have measurable effects on the conformation and conformational stability of either protein. Marked higher stability to temperature and guanidine hydrochloride were observed for thermolysin as compared with neutral protease, as indicated by monitoring conformational transitions with CD measurements at 220 nm. Antisera prepared in rabbits using thermolysin as immunogen do not cross-react with neutral protease, indicating differences of surface structure between the two proteins. On the basis and limitations of the techniques employed, it is proposed that the two sequentially and functionally homologous metalloendopeptidases may have similar conformations at specific regions (active and binding sites, at least) of the polypeptide chain essential for biological function, while some variability in the structure of other regions may be tolerated.

Amino Acids↗
PubMed1979

Lobectomy with bronchoplastic procedures for lung cancer.

From April 1970 to October 1977, 19 patients with lung cancer of the upper lobar bronchus orifice underwent radical lobectomy with major bronchus resection. Sleeve lobectomy was accomplished in 11 cases and wedge lobectomy in the remaining 8. The length of the free bronchial margin in the surgical specimen was less than 1 cm in 3 cases, but limited pulmonary reserve did not allow pneumonectomy. Squamous carcinoma was diagnosed in 14 patients, adenocarcinoma in 2, oat-cell carcinoma in 2, and large cell carcinoma in one. Most cases (70%) were pathological stage I. There was one operative death due to anastomotic leakage (5%), and another patient required pneumonectomy completion. Of 13 patients with non oat-cell carcinoma and adequate bronchial resection, none had local recurrence: 3 patients developed distant metastases, and 10 are alive and disease-free after a follow-up period ranging from 16 to 104 months. The authors conclude that in selected lung cancer patients lobectomy with bronchoplastic procedures is superior to pneumonectomy for tissue sparing advantages.

Adenocarcinoma↗
PubMed1977

Surgical treatment of primary breast cancer.

Radical mastectomy should be replaced by a number of different types of operation which are either more conservative or more extended according to the spread of the disease. For T1N0 cases a new procedure consisting of removal of a quadrant of the breast, with axillary dissection plus radiotherapy on the residual breast tissue, is under evaluation. In T2N0 cases modified radical mastectomy is suggested as a procedure of choice, whilst T3 cases and all N1 cases should still undergo radical mastectomy. Cases with tumor of inner quadrants with positive axillary nodes with benefit from removal of internal mammary nodes. Superradical mastectomy is limited to selected special cases with extensive lymphnode invasion. All N+ cases receive adjuvant chemotherapy (CMF). In locally advanced cases "reductive" surgery may form part of an aggressive chemotherapy programme.

Antineoplastic Agents↗
PubMed1976

Thermal properties of glyceraldehyde 3-phosphate dehydrogenase from Escherichia coli.

The molecular properties of glyceraldehyde 3-phosphate dehydrogenase from E. coli have been evaluated by circular dichroism and fluorescence emission spectroscopy measurements, with the purpose of studying the structural properties which are relevant for a comparison with the enzyme from the obligate thermophile Bacillus stearothermophilus. The enzyme is moderately resistant to heat treatment, being pratically stable when treated for 10 min at 50 degrees C and completely inactivated when heating was performed at 60 degrees C. The secondary structure of the E. coli GPDH appears to be predominatly beta-structure as judged by circular dichroism, showing a negative band centered at about 219 nm. The emission fluorescence of the enzyme shows a maximum at 333 nm upon excitation at 295 nm. In the native E. coli enzyme the tryptophan residues seem to be buried in a hydrophobic region rather than exposed to a polar environment. The structure of the enzyme did not change up to about 50 degrees C, at which temperature thermal inactivation takes place. Upon denaturation the circular dichronic signal at 219 nm gradually decreases, and a red shift of the emission maximum from 333 nm to ca. 345 nm upon heating is indicative that the native structure of the enzyme is unfolded, the tryptophan being exposed to the solvent medium. Since it has been found that the E. coli GPDH closely resembles in many of its properties the B. stearothermophilus enzyme, this bacterial enzyme seems to be useful for comparision with the thermophilic enzyme in studies of its thermostability.

Circular Dichroism↗
PubMed1976

Comparative conformational properties of thermophilic and mesophilic 6-phosphogluconate dehydrogenase.

The structural properties of 6-phosphogluconate dehydrogenase from the mesophilic bacterium E. coli and the thermophilic B. stearothermophilus are compared using circular dichroism and fluorescence emission spectroscopy. The enzymes appear to possess a similar structure which does not change on heating up to the respective temperature of stability of the enzyme. The thermostability of the two 6-phosphogluconate dehydrogenases as determined by activity measurements parallels that determined by CD with the melting profile method, indicating that the loss of biological activity in the enzymes is directly related to the unfolding of the protein molecule. The pattern of unfolding of the proteins by the action of 8 M urea suggests that a core of enhanced conformational stability exists in the B. stearothermophilus enzyme.

Circular Dichroism↗
PubMed1976

[Drug-protein interactions: physico-chemical study on the interactions between chlorpromazine and glutamate dehydrogenase].

The molecular basis of inhibition of glutamate dehydrogenase by chlorpromazine was studied by circular dichoroism, differential spectroscopy and fluorescence. Chlorpromazine appears to induce conformational changes of the enzyme at the position of aromatic amino acids, and tryptophan residues in particular are involved. On the other hand the secondary structure of the enzyme is not influenced by binding of the drug.

Binding Sites↗