KIF13B enhances the endocytosis of LRP1 by recruiting LRP1 to caveolae

Multifunctional low-density lipoprotein (LDL) receptor-related protein 1 (LRP1) recognizes and internalizes a large number of diverse ligands, including LDL and factor VIII. However, little is known about the regulation of LRP1 endocytosis. Here, we show that a microtubule-based motor protein, KIF13B, in an unexpected and unconventional function, enhances caveolin-dependent endocytosis of LRP1. KIF13B was highly expressed in the liver and was localized on the sinusoidal plasma membrane of hepatocytes. KIF13B knockout (KO) mice showed elevated levels of serum cholesterol and factor VIII, and KO MEFs showed decreased uptake of LDL. Exogenous KIF13B, initially localized on the plasma membrane with caveolae, was translocated to the vesicles in the cytoplasm with LRP1 and caveolin-1. KIF13B bound to hDLG1 and utrophin, which, in turn, bound to LRP1 and caveolae, respectively. These linkages were required for the KIF13B-enhanced endocytosis of LRP1. Thus, we propose that KIF13B, working as a scaffold, recruits LRP1 to caveolae via LRP1–hDLG1–KIF13B–utrophin–caveolae linkage and enhances the endocytosis of LRP1.     

Enzymatic desulfurization of dibenzothiophene by a cell-free system of Rhodococcus erythropolis D-1

Abstract Pseudomonas syringae cells were exposed to Cu²⁺ alone or in the precence of acetate, proline or cysteine, at concentrations that reduced free Cu²⁺ to 1/10 of the total copper. Ligand concentrations (designated as isoeffective) were determined experimentally using a Cu²⁺-selective electrode and confirmed by computer calculations using published stability constants. Exposure of P. syringae cells to Cu²⁺ alone resulted in rapid and pronounced cell death, and binding of most of the copper in solution. The addition of acetate, proline or cysteine, a few minutes after Cu²⁺ treatment, resulted in a significant reduction in cell death, and in the amount of copper bound to the cells. For short exposures to Cu²⁺, cysteine was more effective than acetate or proline, but after 60 min of treatment, similar results were observed with these ligands. The addition of ligands before Cu²⁺ resulted in even more reduced copper toxicity. The results showed that, at isoeffective concentrations, weak and moderate copper-ligands can effectively antagonize copper toxicity, and that this protective effect does not require previously equilibrated copper-ligand solutions and is not very dependent of the nature of the ligand.     

Identification of BPESC1, a Novel Gene Disrupted by a Balanced Chromosomal Translocation, t(3;4)(q23;p15.2), in a Patient with BPES

The blepharophimosis syndrome (BPES) is a rare genetic disorder characterized by blepharophimosis, ptosis, epicanthus inversus, and telecanthus. In type I, BPES is associated with female infertility, while in type II, the eyelid defect occurs by itself. The BPES syndrome has been mapped to 3q23. Previously, we constructed a YAC-, PAC-, and cosmid-based physical map surrounding the 3q23 translocation breakpoint of a t(3;4)(q23;p15.2) BPES patient, containing a 110-kb PAC (169-C 10) and a 43-kb cosmid (11-L 10) spanning the breakpoint. In this report, we present the identification of BPESC1 (BPES candidate 1), a novel candidate gene that is disrupted by the translocation on chromosome 3. Cloning of the cDNA has been performed starting from a testis-specific EST, AI032396, found in cosmid 11-L 10. The cDNA sequence of BPESC1 is 3518 bp in size and contains an open reading frame of 351 bp. No significant similarities with known proteins have been found in the sequence databases. BPESC1 contains three exons and spans a genomic fragment of 17.5 kb. Expression of BPESC1 was observed in adult testis tissue. We performed mutation analysis in 28 unrelated familial and sporadic BPES patients, but, apart from the disruption by the translocation, found no other disease-causing mutations. These data make it unlikely that BPESC1 plays a major role in the pathogenesis of BPES.     

Microbiological Asymmetric Hydroxylation of 7-Carboxybicyclo[2.2.1]heptane and hept-2-ene and Their Methyl Esters Giving Potentially Useful Chiral Synthons for Cyclopentanoids

One hundred and nineteen strains of microorganisms (yeasts, bacteria, actinomycetes and fungi) were screened as to the hydroxylation of bicyclo[2.2.1]heptane-7-carboxylic acid, bicyclo[2.2.1]hept-2-ene-7-syn-carboxylic acid, and their methyl esters. Several species belonging to the genera, Bacillus, Streptomyces, Penicillium, Aspergillus, Absidia, Beauveria, Cunninghamella, Drechslera, Mucor and Chaetomium, were found to asymmetrically hydroxylate some or all of the substrates. Bacillus thuringiensis and Aspergillus awamori were the most effective microorganisms for obtaining the chiral products, (lR)-2-hydroxy acids or esters, with enantiomeric purities of 75~90% e.e., which are potential intermediates for (?)-methyl jasmonate or natural prostaglandins.     

Zirex: a Novel Zinc-Regulated Expression System for Lactococcus lactis

Here, we report a new zinc-inducible expression system for Lactococcus lactis, called Zirex, consisting of the pneumococcal repressor SczA and P_czcD. P_czcD tightly regulates the expression of green fluorescent protein in L. lactis. We show the applicability of Zirex together with the nisin-controlled expression system, enabling simultaneous but independent regulation of different genes.     

Studies on the Structure of Gluco-oligosaccharides Obtained from the Partial Acid Hydrolyzate of Sclerotia of Sclerotinia libertiana

The defatted sclerotia powder was partially hydrolyzed with dilute acid, and the material obtained was fractionated by carbon column chromatography, separated into two disaccharides, three trisaccharides and three tetrasaccharides, respectively. In these hydrolyzates α, α-trehalose, Iaminaribiose, and gentiobiose were identified.     

Separation and Purification of Molecular Species of Galactolipids by High Performance Liquid Chromatography

The NaCl concentration of the growth medium affected hydrogen production by Lyngbya sp. (No. 108) strain. Cells grown in medium containing 3% NaCl produced the most hydrogen. The carbohydrate content of this strain also increased with increasing NaCl concentration of the growth medium up to 720 fig/mg cells at 5 % NaCl. In the presence of 20 finlol/ml MFA (monofluoroacetic acid), inhibition of hydrogen production was observed. We extracted the glycogen from this nonheterocystous filamentous cyanobacterium, Lyngbya sp. (No. 108), and observed that glycogen and carbohydrate consumption of this strain is coincident with hydrogen production.

These results led us to the conclusion that the reserve glycogen or other carbohydrate were used as sources of electron donors for hydrogen production, and that the NaCl concentration of the medium affected the hydrogen production by this strain.     

Purification and Some Properties of α-Galactosidase from Tubers of Stachys affinis

An α-galactosidase from tubers of S. affinis was purified about 130 fold by ammonium sulfate fractionation, chromatography on DEAE-cellulose and gel filtration on Sephadex G-75. The purified enzyme showed a single protein band on disc gel electrophoresis. The molecular weight of the enzyme was determined to be approximately 42,000 by gel filtration and 44,000 by SDS disc gel electrophoresis. The optimum reaction pH was 5.2. The enzyme hydrolyzed raffinose more rapidly than planteose. The activation energy of raffinose and planteose by the enzyme was estimated to be 7.89 and 11.4 kcal/mol, respectively. The enzyme activity was inhibited by various galactosides and structural analogs of d-galactose. Besides hydrolytic activity, the enzyme also catalyzed the transfer reaction of d-galactosyl residue from raffinose to methanol.     

Rapid PaperMicrobial Oxygenation of Organo-metallic Bissulfides Leading to Formation of Planar Chirality and C2 Symmetry

1,2-Bis(methylthiomethyl)ferrocene (3) was oxidized by Corynebacterium equi IFO 3730 to give monosulfoxide 4 in two diastereomeric forms with (1S,2R,S_S) and (1S,2R,R_S) configurations in a ratio of 4:1, while 1,1′-bis(methylthiomethyl)ferrocene (5) was oxidized by Penicillium frequentans IFO 5692 to (R)-monosulfoxide 6 and then preferentially to (R,R)-bissulfoxide 7. Thus, the bacterial monooxygenase generated specific planar chirality in the metallocenic monosulfoxide, and the fungal enzyme formed C₂ symmetry in the bissulfoxide.