Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.25.1 (proteasome)
 28,817 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Proteasome subunits are encoded by members of the same gene family and can be divided into two groups based on their similarity to the alpha and beta subunits of the simpler proteasome isolated from Thermoplasma acidophilum. RN3 is the beta-type subunit, N3, of rat proteasomes which has been implicated in the peptidylglutamyl-peptide hydrolase activity of the proteinase complex. We have expressed recombinant RN3 protein in Escherichia coli in order to raise subunit-specific polyclonal antibodies. Identification of the position of RN3 on two-dimensional PAGE gels of purified rat liver proteasomes showed a single protein spot of molecular mass 24 kDa and of pI value of about 5. This protein has a free N-terminus, having undergone post-translational processing. After immunoprecipitation from [35S]methionine-labelled human embryo lung L-132 cells using anti-RN3 antibodies, two radiolabelled spots were observed on two-dimensional PAGE gels, one corresponding to the mature N3, the other of molecular mass 28.5 kDa and pI value around 5, which was probably the unprocessed form of N3. However, the latter protein had a higher molecular mass (31 kDa) than was predicted from the sequence of previously cloned cDNA. Therefore rapid amplification of cDNA ends ("RACE') was carried out to determine the full sequence. The lack of detectable RN3 precursor in purified rat liver proteasomes suggests that the processing probably accompanies assembly of the complex. The half-life of the processing was determined to be 31 min in growing L-132 cells. The unprocessed form of N3 was not observed after immunoprecipitation of 35S-labelled complexes with anti-proteasome antibodies. There was no evidence to suggest that unprocessed N3 is found in precursor complexes which have been implicated in the assembly of some other unprocessed beta-type subunits. Interestingly also, the site of cleavage of N3 (ITR decreases TQN) differs significantly from those of other processed animal beta-type proteasome subunits [(H/T)G decreases TT(T/L)], many of which resemble more closely the cleavage site of the Thermoplasma acidophilum beta subunit.
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PMID:Processing of N3, a mammalian proteasome beta-type subunit. 864 51

In cells, many constituents are able to assemble resulting in large macromolecular machineries possessing very specific biological and physiological functions, e.g. ribosome, spliceosome and proteasome. Assembly of such entities is commonly mediated by transient protein factors. SPAG1 is a multidomain protein, known to participate in the assembly of both the inner and outer dynein arms. These arms are required for the function of sensitive and motile cells. Together with RUVBL1, RUVBL2 and PIH1D2, SPAG1 is a key element of R2SP, a protein complex assisting the quaternary assembly of specific protein clients in a tissue-specific manner and associating with heat shock proteins (HSPs) and regulators. In this study, we have investigated the role of TPR domains of SPAG1 in the recruitment of HSP chaperones by combining biochemical assays, ITC, NMR spectroscopy and molecular dynamics (MD) simulations. First, we propose that only two, out of the three TPR domains, are able to recruit the protein chaperones HSP70 and HSP90. We then focused on one of these TPR domains and elucidated its 3D structure using NMR spectroscopy. Relying on an NMR-driven docking approach and MD simulations, we deciphered its binding interface with the C-terminal tails of both HSP70 and HSP90. Finally, we addressed the biological function of SPAG1 and specifically demonstrated that a SPAG1 sub-fragment, containing a putative P-loop motif, cannot efficiently bind and hydrolyze GTP in vitro Our data challenge the interpretation of SPAG1 possessing GTPase activity. We propose instead that SPAG1 regulates nucleotide hydrolysis activity of the HSP and RUVBL1/2 partners.
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PMID:Binding properties of the quaternary assembly protein SPAG1. 3111 66