5-Aminolevulinate synthase (ALAS) is a mitochondrial enzyme that catalyzes the first step of the heme biosynthetic pathway. The mitochondrial import, as well as the synthesis, of the nonspecific isoform of ALAS (ALAS1) is regulated by heme through a feedback mechanism. A short amino acid sequence, the heme regulatory motif (HRM), is known to be involved in the regulatory function of heme. To determine the role of the HRM in the heme-regulated transport of the nonspecific and erythroid forms of ALAS in vivo, we constructed a series of mutants of rat ALAS1, in which the cysteine residues in the three putative HRMs in the N-terminal region of the enzyme were converted to serine ones by site-directed mutagenesis. The wild-type and mutant enzymes were expressed in quail QT6 fibroblasts through transient transfection, and the mitochondrial import of these enzymes was examined in the presence of hemin. Hemin inhibited the mitochondrial import of wild-type ALAS1, but this inhibition was reversed on the mutation of all three HRMs in the enzyme, indicating that the HRMs are essential for the heme-mediated inhibition of ALAS1 transport in the cell. By contrast, exogenous hemin did not affect the mitochondrial import of the erythroid-specific ALAS isoform (ALAS2) under the same experimental conditions. These results may reflect the difference in the physiological functions of the two ALAS isoforms.

Polarized light scattering from several types of collagen films cast from an acid soluble collagen and solubilized collagens by proteolitic enzyme (except callagenase) and from denatured films under various conditions, was observed under H. and Vv polarization conditions. The scattering patterns from the collagen films were different from those from spherulitic films of polyalphaolefins. The patterns were analyzed in terms of a three-dimentional random assembly model of anisotropic rods resulting in the following conclusions. The polar angle of orientation of scattering elements with respect to the rod axis ranged from 50 to 70°, depending on the type of collagen film. The value of p, which is defined by (al.-a.)/(au-aJ.), changes from very positive in the air-dried state to less positive, zero, or even slightly negative in the swollen state in saline, also depending on the types of collagen film. On the other hand, the characteristic H. patterns were diminished in intensity with denaturation while the Vv patterns-as well as the wide angle X-ray diffraction patternwere not changed to the same extent. This suggests that the light scattering from denatured films arises mainly from correlation in density fluctuations rather than that of orientation of local anisotropy; that is, it is suggested that denaturation causes disintegration of the crystalline superstructure rather than of the crystallites. These crystallites are too small in their correlation distance of the orientation fluctuation to give H. scattering at such small scattering angles as several degrees only. KEY WORDS Polarized Light Scattering I Collagen Film I Denatured Film I Crystalline Texture I Anisotropic Rod I Random Assembly Model/ Calculation I Swelling I Saline I Light scattering from polymer systems in the solid state has been studied from two approaches.

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Depolarized light-scattering intensity distributions from polymer films having a nonspherulitic but so called "rod-like" structure (or "fibrillar" structure) were investigated both theoretically and experimentally. Experimental scattered intensity distributions which were photometrically measured for poly(tetrafluoroethylene) and denatured collagen films were compared with theoretical intensity distributions based upon an assumption of a random assembly of optically anisotropic and homogeneous rods. The results of the comparison show that the theoretical results deviate from the experimental both at small and large scattering angles. The effects of internal disorders of anisotropy and orientation of the optical axes of scattering elements constituting the rods and those of poly-dispersity in the size of the rods are theoretically investigated in order to account for the deviation at large scattering angles. The deviation at small angles may be attributed to an interrod interference effect which is neglected in the theory.

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