Escherichia coli hemolysin (HlyA) is secreted by a specific export machinery which recognizes a topogenic secretion signal located at the C-terminal end of HlyA. This signal sequence has been variously defined as comprising from 27 to about 300 amino acids at the C-terminus of HlyA. We have used here a combined genetic and immunological approach to select for C-terminal HlyA peptides that are still secretion-component. A deletion library of HlyA mutant proteins was generated in vitro by successive degradation of hylA from the 5' end with exonuclease III. Secretion competence was tested by immunoblotting of the supernatant of each clone with an antiserum raised against a C-terminal portion of hemolysin. It was found that the hemolysin secretion system has no apparent size limitation for HlyA proteins over a range from 1024 to 62 amino acids. The smallest autonomously secretable peptide isolated in this selection procedure consists of the C-terminal 62 amino acids of HlyA. This sequence is shared by all secretion-competent, truncated HlyA proteins, which suggests that secretion of the E. coli hemolysin is strictly post-translational. The capacity of the hemolysin secretion machinery was found to be unsaturated by the steady-state level of its natural HlyA substrate and large amounts of truncated HlyA derivatives could still be secreted in addition to full-length HlyA.

Marine industry waste is rich in the extracellular structural protein collagen, excised fragments of which are known to display numerous physiological activities. This study aims at screening waste derived collagen hydrolysates for their utility as peroxide inhibitors in lipid‐based food and cytoprotective agents via cell culture. Collagen was isolated from Pacu and Rohu waste by acid dissolution and salt precipitation. The purified collagen samples were analyzed through electrophoresis, spectral, and elution pattern and confirmed to be of Type‐I. The hydrolysates exhibited a molecular weight around 5 kDa and were found to be in random conformation. The hydrolysates substantially decreased peroxidation levels by 80–90% in linoleic acid model and were equally effective in market available products: cod liver oil and mustard oil. Cell culture assays showed that the hydrolysates were not toxic and capable of increasing cell survival rate by scavenging lipid peroxides generated in situ. Practical applications Peroxidation of lipid‐based food products leads to decreased quality, low benefit‐to‐cost ratio, and imparts harmful effects on consumption, thus requiring synthetic antioxidants to be added to the food. Synthetic oxidation inhibitors currently used carry the risk of carcinogenicity, making the search for natural, nontoxic, and biocompatible peroxide inhibitors a research hotspot. Collagen hydrolysates are bioactive, nontoxic, immune‐compatible, provide added nutritional benefits on consumption and as seen in this study, can be mass isolated from marine industrial wastes making them the perfect functional food additive to be replacing synthetic antioxidants. The utilization of waste would reduce pollution, add substantial value to a common industrial waste and remove the cost of peroxide inhibitor synthesis leading to decrease in the food price and increased shelf life.

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