Mouse and rat cystatin C: Escherichia coli production, characterization and tissue distribution.
Recombinant mouse (Mus musculus) and rat (Rattus norvegicus) cystatin C were produced by expression in Escherichia coli, isolated and functionally characterized. The mouse and rat inhibitors were both fully active in titrations of papain. Determination of equilibrium constants for dissociation (Ki) for their complexes with the target proteinase, cathepsin B, produced values not largely different from that for human cystatin C (Ki 0.07-0.13 nM). Rabbit antisera against mouse and rat cystatin C were produced and used for improved affinity purification of the recombinant inhibitors. Affinity purified immunoglobulins isolated from the antiserum against mouse cystatin C were used for construction of a sensitive enzyme-linked immunosorbent assay. The assay was used to demonstrate a high degree of immunological cross-reactivity between mouse and rat cystatin C and could be used for cystatin C quantification in mouse and rat tissue homogenates. All tissues analyzed contained cystatin C, with a relative content very similar to that of human tissues. For all species, brain tissue contained the highest cystatin C amounts and liver the lowest, whereas kidney, spleen and muscle tissues were intermediate in content. In the mouse, a notable high cystatin C content in parotid gland tissue was observed. The high degree of similarity in distribution pattern and functional properties for mouse, rat and human cystatin C indicates that a murine model should be relevant for studies of the human disease, hereditary cystatin C amyloid angiopathy.
Håkansson
K
K
Department of Clinical Chemistry, University of Lund, University Hospital, Sweden.
Huh
C
C
Grubb
A
A
Karlsson
S
S
Abrahamson
M
M
D000595:Amino Acid Sequence; D000818:Animals; D055316:Cystatin C; D015891:Cystatins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D010206:Papain / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution
C525030:CST3 protein, human; C525031:Cst3 protein, mouse; C525032:Cst3 protein, rat; D055316:Cystatin C; D015891:Cystatins; D011994:Recombinant Proteins; D010206:Papain