Biochimica et Biophysica Acta (BBA) - Specialized Section on Lipids and Related Subjects最新文献

The LDF of hen's egg yolk prepared by a modified method was found to be soluble in water. In this low-density solvent it was possible to separate LDF by flotation into fractions (LDF₁ and LDF₂) owing in part to the maximum effect of the difference in their partial specific volumes. About one-fifth of LDF was estimated to consist of LDF₁, the remainder being LDF₂. Both fractions were polydisperse with molecular weights ranging from 0.5 · 10⁶ to 34 · 10⁶ (mol. wt. 10.3 · 10⁶) for LDF₁ and 0.5 · 10⁶ to 14 · 10⁶ (mol. wt. 3.3 · 10⁶) for LDF₂. Since LDF₁ contained 86.8% and LDF₂ 83.2% lipid, the mean size of their protein moieties was about 1.4 · 10⁶, respectively.

Homogenate of rate testicular tissue (10 000 × g for 20 min) was incubated with labeled lanosterol that was prepared biosynthetically from [2-¹⁴C]mevalonate. The oxidative demethylation of lanosterol to C₂₇ sterols was observed by the rate of release of labeled CO₂. No absolute requirement for added cofactors was demonstrated. The maximal rate of demethylation by testicular tissue was 4 mμmoles/h per 100 mg protein. The supernatant fraction from high-speed centrifugation of testicular tissue or liver supported the same rate of demethylation by liver microsomes. This rate was 6 times greater than the corresponding rate with testicular microsomes. In the presence of mitochondria, labeled CO₂ was liberated from lanosterol by both the demethylation reactions and by the oxidation of the side]hain fragment that is formed during the conversion of C₂₇ sterols to C₂₁ steroids. The rate of conversion of lanosterol to C₂₇ sterols and C₂₁ steroids was determined by difference. Inhibition of demethylation decreased the formation of steroids. The time-course of conversion of labeled lanosterol and [4-¹⁴C]cholesterol to testosterone and the effect of inhibition suggest that cholesterol or other C₂₇ sterols are biosynthetic intermediates between lanosterol and testosterone.

The biosynthesis of lymphatic triglycerides from monoglycerides and acyl-CoA involves two transacylations. The purpose of this paper is to compare the action of the corresponding transacylases (monoglyceride transacylase and diglyceride transacylase) on the two positional isomers of their respective substrates (1- and 2-monoglycerides; 1,2- and 1,3-diglycerides). The enzyme preparation is a microsomal fraction of rat-intestinal mucosa. The experimental conditions are selected in such a way that spontaneous isomerizations of partial glycerides used as substrates or formed during the incubations are substantially avoided. Under these conditions, it is shown that:

• 1.

  1. 2-Monoglycerides from high amounts of 1,2-diglycerides and triglycerides, whereas 1-monoglycerides mainly form 1,3-diglycerides and very low quantities of triglycerides. This observation confirms that monoglyceride transacylase acts in vitro on both isomers of monoglycerides and that 1-monoglyceride acylation mainly occurs on the external carbon. Furthermore, it suggests that 2-monoglycerides formed by pancreatic lipase in the testinal lumen are more efficient acceptors for triglyceride biosynthesis than their isomers 1.

• 2.

  2. The first advantage of 2-monoglycerides over 1-monoglycerides is seen at the level of monoglyceride transacylase itself. Competition experiments show that there exists in rat mucosa a single monoglyceride transacylase acting on both isomers. The maximal velocity reached with both isomers is of the same order for a given concentration of acyl-CoA. But, the affinity of the enzyme and of the complez enzyme-acyl-CoA for 2-monoglycerides is definitely higher. Moreover, the ternary complex enzyme-acyl-CoA-monoglyceride breaks down more rapidly when the monoglyceride is the isomer 2. Since 2-monoglycerides are certainly more abundant in mucosa than 1-monoglycerides, these facts mean that the main reaction catalyzed by monoglyceride transacylase in vivo is the acylation of 2-monoglycerides into 1,2-diglycerides.

• 3.

  3. The second advantage is seen at the level of diglyceride transacylase which is distinct from monoglyceride transacylase in rat mucosa. In the presence of acyl-CoA 0.22 mM, the enzyme forms very little triglycerides from 1,3-diglycerides and high amounts from 1,2-diglycerides.

These results strongly suggest that the main pathway for triglyceride biosynthesis in intestinal mucosa is: 2-monoglycerides → 1,2-diglycerides → triglycerides.

This pathway provides a ready utilization of 2-monoglycerides formed by lipase in intestinal lumen and a junction point at the level of 1,2-diglycerides for the classical pathway starting from α-glycerophosphate. The role played by 2-monoglycerides in the biosynthesis process in vivo is proved in a fully independent way by the fact that most exogenouse