Cardiac troponin T isoforms demonstrate similar effects on mechanical performance in a regulated contractile system

Alteration of troponin T (TnT) isoform expression has been reported in human and animal models of myocardial failure. The two adult beef cardiac TnT isoforms (TnT(3) and TnT(4)) were isolated for comparative functional analysis. Thin filaments were reconstituted containing pure populations of the isoforms. The in vitro motility assay was used to directly compare the effect of the two TnT isoforms on force and unloaded shortening as a function of free calcium. We found no significant differences between the two isoforms in terms of calcium sensitivity, cooperativity, or maximal activation (velocity and force) as assessed in a fully calcium-regulated system. Activation by myosin strong binding was similar for thin filaments containing either of the two TnT isoforms. Whereas maximally activated velocity and cooperativity was depressed at pH 6.5, no difference between thin filaments containing the two isoforms was detected. From the small magnitude of the TnT isoform shifts detected in myocardial failure and the lack of significant mechanical effect detected in the motility assay, variable TnT isoform expression is unlikely to be any functional significance in heart failure.     

Differential sensitivity of two insect GABA-gated chloride channels to dieldrin,fipronil and picrotoxinin

In the central nervous system of both vertebrates and invertebrates inhibitory neurotransmission is mainly achieved through activation of γ-aminobutyric acid (GABA) receptors. Extensive studies have established the structural and pharmacological properties of vertebrate GABA receptors. Although the vast majority of insect GABA-sensitive responses share some properties with vertebrate GABAA receptors, peculiar pharmacological properties of these receptors led us to think that several GABA-gated chloride channels are present in insects. We describe here the pharmacological properties of two GABA receptor subtypes coupled to a chloride channel on dorsal unpaired median (DUM) neurones of the adult male cockroach. Long applications of GABA induce a large biphasic hyperpolarization, consisting of an initial transient hyperpolarization followed by a slow phase of hyperpolarization that is not quickly desensitized. With GABA, the transient hyperpolarization is sensitive to picrotoxinin, fipronil and dieldrin whereas the slow response is insensitive to these insecticides.When GABA is replaced by muscimol and cis-4-aminocrotonic acid (CACA) a biphasic hyperpolarization consisting of an initial transient hyperpolarization followed by a sustained phase is evoked which is blocked by picrotoxinin and fipronil. Exposure to dieldrin decreases only the early phase of the muscimol and CACA-induced biphasic response, suggesting that two GABA-gated chloride channel receptor subtypes are present in DUM neurones. This study describes, for the first time, a dieldrin resistant component different to the dieldrin- and picrotoxinin-resistant receptor found in several insect species.     

Discriminating physiological from non-physiological interfaces in structures of protein complexes: A community-wide study

Reliably scoring and ranking candidate models of protein complexes and assigning their oligomeric state from the structure of the crystal lattice represent outstanding challenges. A community-wide effort was launched to tackle these challenges. The latest resources on protein complexes and interfaces were exploited to derive a benchmark dataset consisting of 1677 homodimer protein crystal structures, including a balanced mix of physiological and non-physiological complexes. The non-physiological complexes in the benchmark were selected to bury a similar or larger interface area than their physiological counterparts, making it more difficult for scoring functions to differentiate between them. Next, 252 functions for scoring protein-protein interfaces previously developed by 13 groups were collected and evaluated for their ability to discriminate between physiological and non-physiological complexes. A simple consensus score generated using the best performing score of each of the 13 groups, and a cross-validated Random Forest (RF) classifier were created. Both approaches showed excellent performance, with an area under the Receiver Operating Characteristic (ROC) curve of 0.93 and 0.94, respectively, outperforming individual scores developed by different groups. Additionally, AlphaFold2 engines recalled the physiological dimers with significantly higher accuracy than the non-physiological set, lending support to the reliability of our benchmark dataset annotations. Optimizing the combined power of interface scoring functions and evaluating it on challenging benchmark datasets appears to be a promising strategy.     

Methylated amino acids in ribosomal proteins from Escherichia coli treated with ethionine and from a mutant lacking methylation of protein L11.

In the present study, the nature, proportions and distribution of methylated amino acids in ribosomal proteins from Escherichia coli grown in the presence of ethionine and from mutant prm 1 were studied. The undermethylated ribosomes had been labeled by addition in vitro or in vivo of radioactive methyl groups from S-adenosylmethionine or from methionine. The following compounds were identified : N alpha-mono-, di- and trimethylalanines, N epsilon-mono-, di- and trimethyllysines, methylamine and N alpha-trimethylalanyllysine. Except for the latter compound and N-alpha-dimethylalanine, all other derivatives had been previously identified in the literature. It is shown that the dipeptide had been in the past mistaken for N epsilon-monomethyllysine, and arises through incomplete hydrolysis in 24 hrs of the N-terminal peptide bond of protein L11. The results of the present study are discussed in the light of previous work on ribosomal protein methylation by the authors and other workers in the field.     

Neurotransmission and sudden infant death. Study of cerebrospinal fluid]

The study of 33 cerebrospinal fluids of infants victims of sudden death shows a very significant increase of the metabolites of dopamine and serotonin. These determinations, compared to a control group, indicate a failure, concerning these two neurotransmitters, which could induce a cardiorespiratory seizure. This failure has likely a multifactorial origin.     

Inter-Alu-like species-specific sequences in the Saccharum complex

Alu sequences constitute the most abundant family of short interspersed nuclear elements, SINEs, in the primate genome. The Alu-PCR method, which consists of amplification between Alu sequences, is usually applied in human genetics to provide polymorphic markers. Here we report the presence of Alu-like sequences in sugarcane and related species by applying the Alu-PCR-like method. Amplifications using a PCR primer defined in conserved regions of Alu human sequences lead to specific complex multiband profiles in all the Saccharum and related genera clones surveyed. The isolation and characterisation of the amplified genus-specific inter-Alu-like fragments allowed us to isolate repeated sequences that are specific for different genera of the Saccharum complex: MsCIR2 from Miscanthus, EaCIR6 and EaCIR7 from Erianthus, and SrCIR2 from Saccharum. Two PCR diagnostic tests were developed from the inter-Alu-like sequences MsCIR2 and EaCIR6, and proved efficient in identifying intergeneric hybrids Saccharum×Miscanthus or Saccharum×Erianthus, respectively. The present study illustrates how the Alu-PCR-like method could help investigating the origin of amphiploid species and monitoring introgression in plants. Received: 7 March 1999 / Accepted: 25 March 1999     

The antitumor properties of the alpha3(IV)-(185-203) peptide from the NC1 domain of type IV collagen (tumstatin) are conformation-dependent

Tumor progression may be controlled by various fragments derived from noncollagenous 1 (NC1) C-terminal domains of type IV collagen. We demonstrated previously that a peptide sequence from the NC1 domain of the alpha3(IV) collagen chain inhibits the in vitro expression of matrix metalloproteinases in human melanoma cells through RGD-independent binding to alpha(v)beta(3) integrin. In the present paper, we demonstrate that in a mouse melanoma model, the NC1 alpha3(IV)-(185-203) peptide inhibits in vivo tumor growth in a conformation-dependent manner. The decrease of tumor growth is the result of an inhibition of cell proliferation and a decrease of cell invasive properties by down-regulation of proteolytic cascades, mainly matrix metalloproteinases and the plasminogen activation system. A shorter peptide comprising the seven N-terminal residues 185-191 (CNYYSNS) shares the same inhibitory profile. The three-dimensional structures of the CNYYSNS and NC1 alpha3(IV)-(185-203) peptides show a beta-turn at the YSNS (188-191) sequence level, which is crucial for biological activity. As well, the homologous MNYYSNS heptapeptide keeps the beta-turn and the inhibitory activity. In contrast, the DNYYSNS heptapeptide, which does not form the beta-turn at the YSNS level, is devoid of inhibitory activity. Structural studies indicate a strong structure-function relationship of the peptides and point to the YSNS turn as necessary for biological activity. These peptides could act as potent and specific antitumor antagonists of alpha(v)beta(3) integrin in melanoma progression.     

The chaperonin GroEL and other heat-shock proteins, besides DnaK, participate in ribosome biogenesis in Escherichia coli

It has been shown that in Escherichia coli the chaperone DnaK is necessary for the late stages of 50S and 30S ribosomal subunit assembly in vivo. Here we focus on the roles of other HSPs (heat-shock proteins), including the chaperonin GroEL, in addition to DnaK, in ribosome biogenesis at high temperature. GroEL is shown to be required for the very late 45S-->50S step in the biogenesis of the large ribosome subunit, but not for 30S assembly. Interestingly, overproduction of GroES/GroEL can partially compensate for a lack of DnaK/DnaJ at 44 degrees C.