Fig. 3.

Experimental workflow for assessment of protein dynamics. Male mice were fed a diet in which 50% of the valine was administered as [²H₈]valine. At different times, pairs of animals were killed, and the seminal vesicle fluid (SV) and sperm preparations (SP) were used for quantitative assessment of the incorporation of labeled valine into protein, by monitoring changes in the labeling pattern of valine-containing peptides. Proteins that are turning over at a high rate would be expected to label rapidly, whereas low turnover proteins would exhibit a gradual shift to stable isotope labeled peptides. A third class of proteins, derived from sperm that are in the process of spermatogenesis during a significant part of the labeling phase, would exhibit delayed labeling if these proteins were only being synthesized only during spermatogenesis and thus demonstrate “lifetime ” kinetics. Sperm proteins that are subject to high turnover even in the epididymis would exhibit the same labeling profile as other high turnover proteins.