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Biomedical subjects

R Kassab

Publications and source records attributed to R Kassab.

At least 73 records · Page 4Linked to original sources
PubMed1984

Muscle gelsolin: isolation from heart tissue and characterization as an integral myofibrillar protein.

A 92-kDa polypeptide present in rabbit and dog cardiac muscle was purified to homogeneity and some of its properties were investigated using biochemical and cytochemical approaches. The protein was found to be similar, if not identical to macrophage gelsolin; it cross-reacts immunologically with anti-rabbit macrophage gelsolin antibody, has a Ca2+-sensitive shortening effect on the actin filaments as judged by the high shear viscometry and sedimentation experiments, and has a similar amino acid composition. In addition, immunoblot and SDS polyacrylamide gel analysis of cardiac muscle extracts obtained at high and low ionic strength showed that this protein is tightly bound to myofibrils, both in the absence and presence of Ca2+, in ventricular as well as in atrial muscle cells. Indirect immunofluorescence microscopy revealed a striated gelsolin staining pattern analogous to that previously observed for the skeletal muscle gelsolin, suggesting that in the muscle cell this protein is sharing the same localisation as actin. Because of its severing and nucleating properties the gelsolin may play a major role in the organization, assembly and turnover of the thin filaments within the muscle cells.

Amino Acids↗
PubMed1984

Recurrent systemic embolic events with valve prosthesis.

Among 1436 patients who underwent valve replacement, the 400 first cases were studied to assess the features of recurrent systemic embolic event. The mean follow-up was 87 months. Three groups of patients were compared: groups A-289 patients without any thromboembolic event (72.25%); group B-78 patients with only one embolic event (19.5%); group C-33 patients with several embolic events (8.25%). The frequency of recurrence was high: one patient out of three (linearized mean 8.9% in group C considering only one recurrence, vs 3.8% in group B). The recurrence have the same location in 45% of patients. The consequences of these embolisms are serious; each event has a 30 to 40% risk of death or major disability. Four variables seem statistically to promote the occurrence of embolic events: mitral prostheses, pre-operative fibrillation, left atrial enlargement, poor anticoagulant therapy. 54 months after the first embolic event, 60% of the patients with poor anticoagulant therapy experience a recurrent thromboembolism vs 20% with adequate therapy. Twenty-six patients of groups B and C had a pathological study of prostheses. Thrombosis of the prostheses was found in 12 out of 18 patients in group B and in 7 out of 8 patients; in group C. Strict observance of anticoagulant therapy is the better way to prevent thromboembolism and especially recurrences. A reoperation is sometimes necessary. Valve re-replacement was performed in 27 cases out of 1436 patients.

Actuarial Analysis↗
PubMed1983

Selective cleavage of the connector segments within the myosin-S1 heavy chain by staphylococcal protease.

The existence of the two connector segments linking the tryptic 50 kDa fragment of skeletal S1 heavy chain to the adjacent 27 kDa and 20 kDa peptides was ascertained by digestion of S1 with staphylococcal protease which was found to act specifically at these particular regions. Three new peptides of Mr 28000, 48000 and 22000 were produced and the novel S1 derivative formed had an intact actin-activated ATPase activity. Amino acid sequence analyses indicated that the 48 kDa and 22 kDa peptides overlap the two connector elements.

Amino Acid Sequence↗
PubMed1983

Structural and actin-binding properties of the trypsin-produced HMM and S1 from gizzard smooth muscle myosin.

The reaction of trypsin on the heavy chain of gizzard myosin and chymotryptic HMM was investigated under restricted fragmentation conditions. The three fragments of the head part with 29 kDa, 50 kDa and 26 kDa were isolated and identified. The 66 K heavy chain segment containing the S1-S2 junction was slowly but extensively degraded liberating a S1-like entity which lacked an intact COOH-terminal 26 kDa region; this isolated species displayed full intrinsic ATPase activities but little actin-binding ability. Tryptic HMM was also formed bearing a fragmented heavy chain and lacking the 20 kDa light chain. Its actin-activated ATPase was derepressed upon cleavage of the 66 kDa segment by papain. We propose that the integral 66 kDa heavy chain component is directly involved in the regulation of the gizzard actomyosin ATPase.

Actins↗
PubMed1983

[Role of surgery in the treatment of chronic aortic insufficiency].

The long-term prognostic factors which determine the indications for surgery are studied on the basis of a series of 198 cases operated for chronic aortic incompetence, in terms of their influence on the actuarial long-term mortality rate due to heart disease and irreversible myocardial dysfunction. The mortality in the first month after surgery is 5 per cent. The factors which significantly modify the long-term outcome of operated patients are clinical (cardiomegaly, degree of preoperative heart failure, ventricular extrasystole, functional grade) and haemodynamic (mean pulmonary artery pressure, end-systolic and-end diastolic left ventricular pressures, left ventricular ejection fraction). Multifactorial studies have shown that ventricular dilatation is the most important prognostic factor. Left ventricular function is severely altered, even in patients with few or no symptoms, but, in the absence of marked functional disturbance, the prognosis is usually good, whatever the repercussions on the myocardium. Surgery should be performed routinely in patients with severe functional disturbance, in the absence of reliable individual criteria capable of predicting when the indications have been exceeded. Surgery is also legitimate in patients with few or no symptoms when investigations reveal severe effects on the left ventricular function.

Adolescent↗
PubMed1981

Structure of the actin-myosin interface.

The topography of the rigor complex between F-actin and myosin heads (S1) has been investigated by carbodiimide zero-length cross-linking. The results demonstrate for the first time that the 95,000-molecular weight (95K) heavy chain of the myosin head enters into van der Waals contact with two neighbouring actin monomers; one is bound to the 50K domain and the other to the 20K domain of the myosin chain. The covalent F-actin-S1 complex can be isolated; it shows a vastly elevated Mg2+-ATPase. Each pair of actin subunits in the thin filament seems to act as a functional unit for specific binding of a myosin head and stimulation of its Mg2+-ATPase activity.

Actins↗
PubMed1981

Structural aspects of actomyosin interaction.

Actin binding to myosin-S1 modulates the limited tryptic cleavage of the COOH-terminal region of the 95K heavy chain at the joint connecting the 75K and 20K peptide units; concomitantly actin affords total protection against the resulting loss of acto-S1 Mg2+-ATPase activity. The specificity of the actin effect is illustrated by the fact that it exerts itself not only on free S1 but also on the intact myosin molecule. Mg2+-ATP and Mg2+-ADP impair the protective action of actin to an extent closely related to their respective affinity for the acto-S1 complex. Tryptic fragmentation of S1 heavy chain under highly controlled conditions, using trypsin to S1 weight ratios in the range 1:1000 - 1:1500 led us to establish that peptide bond cleavage at the 75K-20K junction is a sequential process giving rise first to a 22K peptide intermediate which is subsequently converted to the stable 20K fragment. Most importantly, it is also demonstrated that the loss of S1 activation by actin is not due to the initial scission of the 75K-22K linkage but is intimately associated with the breakdown of the 22K precursor into its 20K moiety. Three trypsin-modified S1 derivatives, the heavy chain of which is a complex of two or three fragments, were purified. A detailed analysis of the C-termini of these fragments, as compared to the C-terminal structure of the intact heavy chain, indicated that the 20K fragment is formed mainly through the degradation of a NH2-terminal 2K segment in the 22K precursor and that this proteolytic event is the only one accounting for the acto-S1 ATPase loss. Cross-linking experiments exploiting the reaction of a carbodiimide reagent with rigor complexes containing either fluorescent actin or fluorescent fragmented S1 revealed unequivocally the attachment of the actin monomer to recognition sites on the 20K and 50K units of S1 heavy chain. Specific interactions between the C-terminal 20K domain and light chain LC2 are proposed as being part of the molecular mechanism of the myosin-linked regulation of actomyosin interaction.

Actins↗
PubMed1981

Primary structure of lobster-muscle arginine kinase. Amino and carboxyl-terminal structure of the enzyme and complete alignment of the cyanogen-bromide peptides.

The acetylation state of the blocked NH2-terminal end of arginine kinase was characterized by the occurrence of 1 mol acetyl group/mol protein; a 5-residue segment corresponding to the amino-terminal portion was isolated from a pronase digest of the enzyme and its amino acid sequence determined as N-acetyl Ala-Asx-Ala-Thr. Arginine kinase is terminated at the carboxyl end by the sequence Lys-GluMetOH; this particular 3-residue sequence is repeated three times in the overall structure of the protein and is present in three CNBr fragments. One of these, a peptide of 14 amino acid residues, was identified in the course of this study and its amino acid sequence determined. Its location at the COHO-terminal end of the enzyme was recognized on the basis of investigations carried out with des-MetOH-Glu-arginine kinase, a specific proteolytic derivative. The alignment of the eight CNBr-fragments which constitute the arginine kinase molecule was established according to the sequential and compositional properties of seven unique tryptic methionyl peptides isolated from the whole protein. The alignment was confirmed by using BNPS-skatole fragments of the enzyme as another protein source.

Acetylation↗
PubMed1980

Photoaffinity labelling of arginine kinase and creatine kinase with a gamma-P-substituted arylazido analogue of ATP.

1. An ATP analogue with a photoactivated azide group attached to the gamma-phosphate via an amide bond, ATP gamma-p-azidoanilide, appeared to have potential use as a photoaffinity label for the nucleotide-binding regions of ATP: guanidine phosphotransferases. Upon photolysis in the presence of lobster muscle arginine kinase and rabbit muscle creatine kinase, the analogue is converted to a potent inhibito of these two kinases. This photo-dependent inhibition is specific as it cannot be induced by azidoaniline, a mixture of azidoaniline and ATP or by ATP gamma-p-aminoanilide. Preirradiated under suitable conditions, the photoanalogue still shows a transitory inhibitory effect which, however, slowly vanishes with time (t0.5 = 3 h). 2. The photoinhibition is significantly decreased by the presence of ATP or ADP but is completely prevented by the addition of a mixture of nucleotide and guanidine substrates. Differential spectroscopy and affinity chromatography on Sepharose-ATP demonstrated the inability of photoinactivated arginine kinase and creatine kinase to recognize their nucleotide substrates. 3. Experiments with [14C]ATP gamma-p-azidoanilide indicated that photolysis is associated with an irreversible and stoichiometric binding of the ATP analogue to the enzymes. Autoradiographs made with the peptide maps corresponding to the tryptic digests of each 14C-labelled photomodified enzyme showed an unexpected highly specific labelling of the proteins. 4. Thiiol titrations of the kinases which have been subjected to various photolysis conditions led to the conclusion that the arylnitrene moiety of the photoanalogue is covalently attached to the single reactive cysteinyl side chain present in the active-site region of the two homologous kinases. This amino acid residue appears, therefore, to be located near the phosphate chain binding subsite occupied by the ATP analogue and probably also by the natural nucleotide substrates.

Adenosine Triphosphate↗
PubMed1979

Involvement of an arginyl residue in the catalytic activity of myosin heads.

1. Phenylglyoxal reacts rapidly with isolated myosin heads (subfragment 1) and induces two successive and distinguishable effects on their enzymic properties: first, a twofold activation of the Ca2+ and Mg2+-dependent ATPases with no effect onthe K+-ATPase followed by inhibition of the K+, Ca2+ and actin-activated Mg2+-ATPases. A specific protein-reagent reagent complex is formed during the second phase of the modification reaction (Ki approximately 5 x 10(-3) M). 2. ADP and ATP with or without cations provide efficient protection only against the loss of ATPase activities, suggesting that the second inhibitory process is occurring at or close to the active site. 3. On the basis of [14C]phenylglyoxal-labelling experiments and the composition of modified subfragment-1 derivatives, it is demonstrated that the sequential modification of two reactive arginyl residues is responsible for the observed activation-inhibition phenomena. Blocking of the first reactive residue produces a shift in the pH/activity curves related to the Ca2+ and Mg2+-dependent ATPases with an apparent activation effect. Modification of the second guanidino group does not destroy the affinity of the protein for the nucleotide substrates but does alter the nucleotide binding site as reflected in the inability of Mg2+. ATP to dissociate the modified subfragment-1--actin complex. It is concluded that electrostatic interactions between this positively charged group and the negatively charged ATP and ADP molecules may be critical for the hydrolytic efficiency of myosin heads. 4. After dissociation and separation of the polypeptide constituents of the protein in acetic acid medium, both labelled sites are found to reside in the heavy chain.

Adenosine Triphosphatases↗
PubMed1977

Amino acid sequence of a cyanogen bromide fragment containing the two tryptophanyl residues of lobster arginine kinase (Homarus vulgaris).

Lobster arginine kinase [EC 2.7.3.3] contains 2 tryptophanyl residues and 9 methionyl residues. The whole carboxymethylated protein was first subjected to CNBr cleavage and the resulting fragments were isolated by gel filtration and other experimental approaches. One fragment, CB5, which contains 60 residues including the two tryptophanyl residues and two of the five cysteinyl residues of the protein, was characterized and the results are reported inthis paper. The overall strategy for the establishment of the complete sequence of this fragment was based on the use of three types of peptides: (a) whole cyanogen bromide peptide CB5 which was partially characterized by automatic Edman degradation using a sequencer: 42 steps were performed out of 60 residues, (b) tryptic peptides of CB5, (c) peptides formed by cleavage of S-carboxymethylated arginine kinase (whole protein) at the two tryptophanyl residues with BNPS-skatole. The complete amino acid sequence of the CNBr polypeptide (CB5) which contains the two tryptophanyl residues of the whole protein was established.

Amino Acid Sequence↗