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The Synthesis of Hydroxyproline 

 within Osteoblasts 



[Abstract] 



SYLVIA FITTON JACKSON 



Medical Research Council Biophysics Research Unit 

 W heatstone Laboratory , King's College, London 



Biochemical and morphological methods are being used to study the stages 

 of synthesis of intercellular material in active collagen-producing tissue cultures. 

 The direct oxidation of proline already bound in peptide linkage may be an 

 important step in the sequence of the synthetic processes which lead to the 

 formation of collagen protein [Stetten, 1949]. In previous work it has been 

 found that appreciable amounts of protein-bound hydroxyproline were formed 

 during the first 24 hr of culture before the appearance of characteristic collagen 

 fibrils [Fitton Jackson and Smith, 1957]. Free C 14 -L-proline was also readily 

 incorporated into the proteins of the growing tissue, and as much as 20 per 

 cent was converted to protein-bound C 14 -hydroxyproline [Smith and Fitton 

 Jackson, 1957]. 



Cell fractionation studies have been made on similar tissue cultures in an 

 attempt to establish whether the site of incorporation of free proline into the 

 proteins of the cell was the same as that of the formation of the protein-bound 

 hydroxyproline. The cultures were grown in contact with C 14 -L-proline for 

 various times and subsequently homogenized and subjected to differential cen- 

 trifugation in 0.88 M sucrose solution. The amount of labeled proline incor- 

 porated and converted to hydroxyproline in the six isolated fractions was meas- 

 ured. Observations were made in parallel on the morphology of the whole 

 cells and the various cellular fractions by means of the electron microscope. 



Chemical analyses demonstrated the consistent presence of protein-bound 

 hydroxyproline in the fractions of larger particle size (3000 A). The results 



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