Purification and properties of parathyroid hormone-related peptide isolated from milk.
The occurrence and properties of PTH-related peptide (PTH-RP) in milk was investigated. PTH-RP was purified to homogeneity from human and bovine milk using heat and acid to precipitate milk proteins followed by ion exchange chromatography and reverse-phase HPLC. The peak of PTH-RP from HPLC was detected using a sensitive bone cell bioassay. A single band of peptide was detected on silver-stained polyacrylamide gels, which migrated as a 20-21-kDa macromolecule. PTH-RP isolated from either human or bovine milk had similar electrophoretic mobilities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The partially purified bovine PTH-RP stimulated cAMP production in UMR106-01 and OK cell lines and elicited a concentration-dependent inhibition of sodium-dependent phosphate transport in OK cells. Incubation of milk extracts with an anti-PTH antiserum did not affect their bioactivity, whereas an antihuman PTH-RP 1-34 antiserum markedly reduced the cAMP response of UMR106-01 cells to the immunoabsorbed milk extracts. A PTH antagonist, norleu PTH 3-34, blocked the stimulation of cAMP production in UMR106-01 cells treated with milk extracts. PTH-RP immunoreactivity and bioactivity occurred in milk extracts of diverse animals from both eutherian and metatherian (marsupial) species. Porcine colostrum also had immunoreactive PTH-RP, although the levels were lower than the immunoreactive PTH-RP concentrations observed in milk samples collected at 7 and 14 days of lactation. Thus, a 20-21-KDa PTH-RP is secreted into milk where it could play a role in the development of suckling, newborn animals.
Thurston
A W
AW
Department of Pharmacology, Missouri University, Columbia 65212.
Cole
J A
JA
Hillman
L S
LS
Im
J H
JH
Thorne
P K
PK
Krause
W J
WJ
Jones
J R
JR
Eber
S L
SL
Forte
L R
LR
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001681:Biological Assay; D015153:Blotting, Western; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003126:Colostrum / Q000032:analysis; D000242:Cyclic AMP / Q000096:biosynthesis; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D006041:Goats; D006801:Humans; D008892:Milk / Q000032:analysis*; D008895:Milk, Human / Q000032:analysis*; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism; D044162:Parathyroid Hormone-Related Protein; D010710:Phosphates / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification* / Q000494:pharmacology; D013552:Swine; D014407:Tumor Cells, Cultured
C470121:PTHLH protein, human; D044162:Parathyroid Hormone-Related Protein; D010710:Phosphates; D011506:Proteins; D000242:Cyclic AMP; D000262:Adenylyl Cyclases