Abstract
The sheep meat industry in New Zealand and Australia is huge, but produces a lot of ined-
ible coproducts, including organs like lungs. These organs carry out a range of biochem-
ical functions, requiring proteins with a range of distinct properties. There is untapped
potential to increase the value of those coproducts by extracting proteins and protein hy-
drolysates with useful bioactive or material properties, for example antioxidant or gelling
abilities. Lungs contain a rich environment of proteins like surfactants, collagens, elastins
and immunoglobulins to extract. Hence, this project investigates the extraction of protein
and peptide fragments from sheep lungs.
Sheep lung was found to be easily homogenised and separated into soluble and insolu-
ble components, although there was an intermediary layer that contaminated the insoluble
fraction, possibly caused by pulmonary surfactants. This layer was removed with repeated
NaOH washes. Proximate analysis of homogenised whole sheep lungs confirmed that the
majority of them (86%) is protein by dry weight, at least for caudal samples. The soluble
fraction resulted in five clear peak fractions after size exclusion chromatography. Mass
spectrometry found that the majority of the protein in the soluble fraction consisted of
blood proteins. A bioinformatic pipeline was developed to find orthologues and annotate
the ovine proteins discovered with mass spectrometry. This found bovine and human or-
thologues for 99% and 95% of the discovered proteins, respectively, leading to 85% more
annotations than with ovine annotations alone.. Over 15% of the distinct proteins were
noted as blood proteins, and 8%–13% noted as a lung protein. The proteins discovered
include antioxidants and antimicrobial proteins that are enriched in lungs.
The insoluble portion of whole lung homogenate was subjected to pH shifts, high
temperatures, and enzymatic digestion to solubilise as many protein species and peptide
chains as possible. Protein solubility increased markedly at basic pH (pH 10), and acidic
pH also appeared to solubilise additional proteins and hydrolyse existing ones. Protein
in the soluble portion of homogenate appeared to decrease in solubility with a decrease
in pH. Base-acid-base pH shift is an inexpensive early purification step. Acid precipi-
tation was especially effective at isolating two proteins, possibly albumin and a globin.
Enzyme digestions on NaOH-washed insoluble homogenate with bromelain, papain and
PR6L found specific hydrolysis products around 18 kDa–20 kDa for bromelain and papain,
as well as high weight bands for PR6L. Digestion with pepsin intermittently solubilised dis-
tinct banding patterns characteristic of collagen. Subjecting the homogenate to 90 °C for
19 hours resulted in non-specific degradation and aggregation of soluble protein, making
it unsuitable for the goals of this project. For the NaOH-washed insoluble-only material,
specific solubilisation products were observed after an hour at 80 °C.