We report herein the cDNA cloning of a novel rat acyl-CoA synthetase (ACS) that preferentially uses arachidonate and eicosapentaenoate. This newly identified ACS (designated ACS4) contains 670 amino acids and is 68% identical to rat ACS3, a previously characterized ACS that is highly expressed in brain. ACS4 was overproduced in Escherichia coli and the resulting enzyme was purified to homogeneity. The purified enzyme utilizes arachidonate and eicosapentaenoate most preferentially among C 8 -C 22 saturated fatty acids and C 14 -C 22 unsaturated fatty acids. Kinetic analyses revealed that the enzyme has a high affinity for arachidonate and eicosapentaenoate and low affinity for palmitate. ACS4 transcripts are detectable in a wide range of tissues, with the highest level in adrenal gland. Immunoreactivity to ACS4 was detected in the zona fasciculata and reticularis of adrenal gland, in the corpus luteum and stromal luteinized cells in ovary, and in the Leydig cells of testis.

Isolation and characterization of a rat brain cDNA identified a third acyl-CoA synthetase (ACS) designated ACS3. The deduced amino acid sequence of the cDNA revealed that ACS3 consists of 720 amino acids and exhibits a structural architecture common to ACSs from various origins. ACS3 expressed in COS cells was purified to near homogeneity. The purified ACS3 resolved by SDS-polyacrylamide gel electrophoresis into two major proteins of 79 and 80 kDa. Cell-free translation of a synthetic mRNA encoding the entire region of ACS3 revealed that the two isoforms were derived from the same mRNA. The purified ACS3 utilizes laurate and myristate most efficiently among C8-C22 saturated fatty acids and arachidonate and eicosapentaenoate among C16-C20 unsaturated fatty acids. Northern blot analysis revealed that ACS3 mRNA is most abundant in brain and, to a much lesser extent, in lung, adrenal gland, kidney, and small intestine. During the development of the rat brain, expression of ACS3 mRNA reached a maximum level at 15 days after birth and then declined gradually to 10% of the maximum in the adult brain.

Isolation and characterization of a human cDNA demonstrated a novel lipoprotein receptor designated apolipoprotein E receptor 2 (apoER2). The new receptor consists of five functional domains resembling the low density lipoprotein (LDL) and very low density lipoprotein (VLDL) receptors. LDL receptor deficient Chinese hamster ovary cells expressing human apoER2 bound apoE rich ␤-migrating VLDL with high affinity and internalized. LDL was bound with much lower affinity to these cells. The 4.5-and 8.5-kb mRNAs for the receptor were most highly expressed in human brain and placenta. In rabbit tissues, multiple species of the mRNA with 4, 4.5, 5.5, 8.5, and 11 kb were detected most intensely in brain and testis and, to a much lesser extent, in ovary, but were undetectable in other tissues. In rat adrenal pheochromocytoma PC12 cells, the receptor mRNA was induced by treatment of the cells with nerve growth factor. The receptor transcripts were detectable most intensely in the cerebellar cortex, choroid plexus, ependyma, hippocampus, olfactory bulb and, to a much lesser extent, in the cerebral cortex as revealed by in situ hybridization histochemistry. In the cerebellar cortex, the receptor transcripts were densely deposited in Purkinje cell somata.Receptor-mediated endocytosis of plasma lipoproteins plays an important role in the metabolism of cholesterol and triglyceride in the body. The low density lipoprotein (LDL) 1 receptor, one of the best characterized cell surface receptors, mediates cholesterol homeostasis in the body (1). The LDL receptor binds apolipoprotein B-100 containing LDL and apolipoprotein E (apoE)-containing lipoproteins, whereas the recently found very low density lipoprotein (VLDL) receptor binds only apoEcontaining lipoproteins (2, 3). Both the LDL receptor (4 -7) and VLDL receptor (2, 8 -14) consist of five functional domains: (i) an amino-terminal ligand binding domain composed of multiple cysteine-rich repeats; (ii) an epidermal growth factor (EGF) precursor homology domain, which mediates the acid-dependent dissociation of the ligands from the LDL receptor (15); (iii) an O-linked sugar domain; (iv) a transmembrane domain; and (v) a cytoplasmic domain with a coated pit targeting signal (16). Genetic deficiencies of the LDL receptor give rise to familial hypercholesterolemia, one of the most common genetic diseases in humans (17). Mutations in the chicken VLDL receptor gene lead to the failure to produce offspring (13, 18).Lipoprotein metabolism in the central nervous system (CNS) has been poorly understood, despite the importance of lipids in some specialized neural membranes, such as myelin. Most of lipids in the CNS are actively synthesized in the CNS itself and deposited in large amounts during the early phase of development (19, 20). The rate of cholesterol and fatty acid synthesis in the brain is high during the myelinating period and declines thereafter (19,20). Although most of lipids in the brain are believed to be synthesized within the brain itself, small amounts of cholesterol (21)...

We report here the identification, characterization, and expression of a novel rat acyl-CoA synthetase (ACS) designated as ACS5. ACS5 consists of 683 amino acids and is approximately 60% identical to the previously characterized ACS1 and ACS2. ACS5 was overproduced in Escherichia coli cells and then purified to near homogeneity. The purified enzyme utilized a wide range of saturated fatty acids similar to those utilized by ACS1 and ACS2, but differed in its preference for C16-C18 unsaturated fatty acids. Northern blot analysis revealed that ACS5 mRNA is present most abundantly in the small intestine, and to a much lesser extent in the lung, liver, adrenal gland, adipose tissue, and kidney. In situ hybridization of rat ileum revealed abundant accumulation of ACS5 transcripts in foveolar epithelial cells. The hepatic level of ACS5 mRNA was significantly increased by refeeding a fat-free high sucrose diet and reduced by fasting or refeeding a high cholesterol diet, whereas that in the small intestine was not significantly altered by various dietary conditions. In contrast to the absence of ACS1 mRNA in undifferentiated 3T3-L1 preadipocytes, ACS5 mRNA was present in proliferating 3T3-L1 preadipocytes and its level remained unaltered during differentiation, suggesting that ACS5 may provide the acyl-CoA utilized for the synthesis of cellular lipids in proliferating preadipocytes.

The gene for the human substance P receptor (NK-1) was cloned using cDNA probes made by the polymerase chain reaction from primers based on the rat sequence. The gene spans 45-60 kb and is contained in five exons, with introns interrupting at sites homologous to those in the NK-2 receptor gene. Analysis of restriction digests of genomic DNA from mouse/human cell hybrids indicates the NK-1 receptor is a single-copy gene located on human chromosome 2. Polymerase chain reaction using primers based on the 5' and 3' ends of the coding sequence was used to generate full-length cDNAs from human lung and from IM9 lymphoblast cells. When transfected into COS-7 cells, the NK-1 receptor binds 125I-BHSP with a Kd of 0.35 +/- 0.07 nM and mediates substance P induced phosphatidylinositol metabolism. The receptor is selective for substance P; the relative affinity for neurokinin A and neurokinin B is 100- and 500-fold lower, respectively. Human IM9 lymphoblast cells express relatively high levels of the NK-1 receptor, and Northern blot analysis indicates modulation of mRNA levels by glucocorticoids and growth factors, suggesting that this cell line may be useful as a model for studying the control of NK-1 receptor gene expression.

We have prepared meso-bis(diarylamino)-substituted porphyrins (cis-ZnP and trans-ZnP), meso-diarylaminosubstituted porphyrin (mono-ZnP), and porphyrin reference without the meso-diarylamino group (ZnP) to evaluate the effects of substituent number and position of the diarylamino groups on the optical, electrochemical, and photovoltaic properties of the porphyrins for the first time. With increasing the number of the diarylamino groups, the light-harvesting properties were improved in the visible region. The optical and electrochemical HOMO-LUMO gaps were parallel to those estimated by DFT calculations. The photovoltaic properties were compared under the optimized conditions in which a sealed device structure with TiCl 4 -treated, TiO 2 double layers was used. The power conversion efficiency (η) was in the order of the trans-ZnP (3.8%) < ZnP (4.4%) < cis-ZnP (5.5%) < mono-ZnP (6.5%) sensitized TiO 2 cells. The low light-harvesting efficiency (LHE) rather than the high adsorbed photon-to-current efficiency (APCE) in the ZnP-sensitized solar cell leads to the moderate short circuit current (J SC ), whereas both the high APCE value and the moderate LHE value in the mono-ZnP-sensitized TiO 2 cell result in the highest J SC value. Considering the similarity in the LHE values of the trans-ZnP and cis-ZnP-sensitized cells, the APCE values correlate the J SC values. The open circuit potentials (V OC ) of the trans-ZnP and cis-ZnP-sensitized TiO 2 cells are lower than those of the mono-ZnP and ZnP-sensitized TiO 2 cells. Overall, the difference in the J SC and V OC values of the four TiO 2 cells leads to that in the η values. The basic information obtained here will be useful to design molecular structures of porphyrins for highly efficient dye-sensitized solar cells.

Novel benzo[f]quinoxalino [2,3-β]porphyrin carboxylic acid (ZnBQA) and cyanoquinoxalino [6,7-β]porphyrin carboxylic acid (ZnQCA) have been synthesized to evaluate the effects of the π-elongation and the fused position of quinoxaline-fused porphyrins on the optical, electrochemical, and photovoltaic properties. ZnBQA showed a split, red-shifted Soret band relative to that of quinoxalino[2,3-β]porphyrin acid (ZnQMA), while the Q bands are rather blue-shifted. On the other hand, both Soret and Q-bands of ZnQCA are red-shifted compared to those of ZnQMA. The optical HOMO-LUMO gaps are consistent with those estimated by density functional theory calculations. The photovoltaic properties were compared under the optimized conditions in which a sealed device structure with TiCl 4 -treated, TiO 2 double layers was used. The ZnBQA cell exhibited a relatively high power conversion efficiency (η) of 5.1%, while the ZnQCA cell yielded a low η value of 0.80%. Both of the η values are smaller than those of reference cells under the optimized conditions (η ) 6.3% for ZnQMA; η ) 8.4% for N719). The weak electronic coupling between the LUMO of ZnBQA and a conduction band (CB) of the TiO 2 may result in the low electron injection efficiency as well as the low incident photon-to-current efficiency (IPCE) for the ZnBQA cell (maximum IPCE ) 60%) relative to the ZnQMA cell (maximum IPCE ) 75%), leading to the lower η value of the ZnBQA cell than that of the ZnQMA cell. The ZnQCA cell exhibited the further low IPCE value up to 10% due to the short fluorescence lifetimes (0.2, 5 ps) that are comparable to the typical time scale (0.1-10 ps) of electron injection processes from a porphyrin excited singlet state to a TiO 2 surface. In addition, the open circuit potential of the ZnQCA cell also significantly decreased by the effect of the charge recombination from the injected electrons in the CB of the TiO 2 to the I 3 derived from the loose packing of ZnQCA molecules on the TiO 2 surface.

We have prepared a novel pushÀpull porphyrin with an electrondonating diarylamino group at the meso-position and an electron-withdrawing 4-carboxy-2,3,5,6-tetrafluorophenylethynyl anchoring group at the opposite mesoposition to address the substitution effects of the electron-withdrawing fluorine atoms on photovoltaic properties for the first time. The fluoro-substituted porphyrin showed slightly improved light-harvesting properties when compared to the reference porphyrin in solution, due to the enhancement of charge-transfer character. The fluoro-substituted porphyrin-sensitized TiO 2 cell exhibited a moderate power conversion efficiency of 4.6% but was lower than the corresponding value (6.9%) of the reference porphyrin-sensitized TiO 2 cell. The carboxylic group in the reference porphyrin binds to the TiO 2 surface with bidentate coordination, leading to formation of the densely packed monolayer on the TiO 2 surface, whereas both of the electronwithdrawing fluorine atoms and the carboxylic group of the fluoro-substituted porphyrin interact with the TiO 2 surface, adopting relatively parallel orientation to the TiO 2 surface with monodentate binding of the carboxylic acid to the TiO 2 , eventually yielding to the small surface coverage and low cell performance due to the fast charge recombination and small amount of the long-lived porphyrin radical cation. Such fundamental information may be useful for the molecular design of highly efficient dye-sensitized solar cells based on pushÀpull porphyrins.