Bacteria use trans-translation and the alternative rescue factors ArfA (P36675) and ArfB (Q9A8Y3) to hydrolyze peptidyl-tRNA on ribosomes that stall near the 3' end of an mRNA during protein synthesis. The eukaryotic protein ICT1 (Q14197) is homologous to ArfB. In vitro ribosome rescue assays of human ICT1 and Caulobacter crescentus ArfB showed that these proteins have the same activity and sub...

Rare AGA or AGG codons close to the initiation codon inhibit protein synthesis by a tRNA-sequestering mechanism as toxic minigenes do. To further understand this mechanism, a parallel analysis of protein synthesis and peptidyl-tRNA accumulation was performed using both a set of lacZ constructs where AGAAGA codons were moved codon by codon from +2, +3 up to +7, +8 positions and a series of 3-8 c...

The dependence of the Vmax and Km on the length of the peptide moiety in the peptidyl-tRNA series (Gly)n-Val tRNA, was measured in the system peptidyl-tRNA hydrolase-peptidyl-tRNA. It was found that the Km value decreases from 7.2 X 10-7 M for Gly-Val-tRNA to 4.6 X 10-7 M FOR (Gly)2-Val-tRNA and to 1.7 X 10-7M for (Gly)3-Val-tRNA; further increase of the peptide chain is not followed by decrea...

The ribosome is the molecular machine responsible for protein synthesis in all living organisms. Its catalytic core, the peptidyl transferase center (PTC), is built of rRNA, although several proteins reach close to the inner rRNA shell. In the Escherichia coli ribosome, the flexible N-terminal tail of the ribosomal protein L27 contacts the A- and P-site tRNA. Based on computer simulations of th...

Peptide bond formation is the main catalytic function of the ribosome. The mechanism of catalysis is presumed to be highly conserved in all organisms. We tested the conservation by comparing mechanistic features of the peptidyl transfer reaction on ribosomes from Escherichia coli and the Gram-positive bacterium Mycobacterium smegmatis. In both cases, the major contribution to catalysis was the ...

Protein L27 has been implicated as a constituent of the peptidyl transferase center of the Escherichia coli 50 S ribosomal subunit by a variety of experimental observations. To define better the functional role of this protein, we constructed a strain in which the rpmA gene, which encodes L27, was replaced by a kanamycin resistance marker. The deletion mutant grows five to six times slower than...

The chloramphenicol (Cm)-inducible cmlA gene of Tn1696 specifies nonenzymatic resistance to Cm and is regulated by attenuation. The first eight codons of the leader specify a peptide that inhibits peptidyl transferase in vitro. Functionally similar, but less inhibitory, peptides are encoded by the leaders of Cm-inducible cat genes. However, the cat and cmlA coding sequences are unrelated and sp...

Purified peptidyl dipeptidase (angiotensin I converting enzyme or kininase II) from human lung or hog kidney is inhibited by commercially prepared plasma protein preparations, by human serum albumin and by the additive albumin stabilizer, acetyltryptophan. After the initial steps of purification, albumin was detected by immunodiffusion as a component in human lung peptidyl dipeptidase preparati...

Acidified sodium chlorite cleaves isodityrosine and solubilizes covalently bound hydroxyproline-rich material from cell walls. This has been taken as evidence that isodityrosine acts as a cross-link holding the hydroxyproline-rich glycoprotein extensin in the cell wall. However, acidified chlorite was found to cleave peptide bonds in salt-soluble extensin and in bovine serum albumin (BSA). This...

alpha-Amidation is catalyzed by two enzymatic activities, peptidyl-glycine alpha-hydroxylating mono-oxygenase (PHM) and peptidyl-alpha-hydroxyglycine alpha-amidating lyase (PAL), denoted collectively as peptidyl-glycine alpha-amidating mono-oxygenase (PAM), which also may include transmembrane and cytoplasmic domains. PAM is present in mammalian pancreas, where it appears to be abundant in the ...