Differential mechanisms of retinoid transfer from cellular retinol binding proteins types I and II to phospholipid membranes

Cellular retinol-binding proteins types I and II (CRBP-I and CRBP-II) are known to differentially facilitate retinoid metabolism by several membrane-associated enzymes. The mechanism of ligand transfer to phospholipid small unilamellar vesicles was compared in order to determine whether differences in ligand trafficking properties could underlie these functional differences. Unidirectional transfer of retinol from the CRBPs to membranes was monitored by following the increase in intrinsic protein fluorescence that occurs upon ligand dissociation. The results showed that ligand transfer of retinol from CRBP-I was >5-fold faster than transfer from CRBP-II. For both proteins, transfer of the other naturally occurring retinoid, retinaldehyde, was 4-5-fold faster than transfer of retinol. Rates of ligand transfer from CRBP-I to small unilamellar vesicles increased with increasing concentration of acceptor membrane and with the incorporation of the anionic lipids cardiolipin or phosphatidylserine into membranes. In contrast, transfer from CRBP-II was unaffected by either membrane concentration or composition. Preincubation of anionic vesicles with CRBP-I was able to prevent cytochrome c, a peripheral membrane protein, from binding, whereas CRBP-II was ineffective. In addition, monolayer exclusion experiments demonstrated differences in the rate and magnitude of the CRBP interactions with phospholipid membranes. These results suggest that the mechanisms of ligand transfer from CRBP-I and CRBP-II to membranes are markedly different as follows: transfer from CRBP-I may involve and require effective collisional interactions with membranes, whereas a diffusional process primarily mediates transfer from CRBP-II. These differences may help account for their distinct functional roles in the modulation of intracellular retinoid metabolism.     

Effect of apolipoprotein A-IV genotype and dietary fat on cholesterol absorption in humans

We investigated the effect of the A-IV-2 allele, which encodes a Q360H substitution in apolipoprotein (apo) A-IV, and dietary fat on cholesterol absorption in humans. In three separate studies we compared fractional intestinal cholesterol absorption between groups of subjects heterozygous for the A-IV-2 allele (1/2) and homozygous for the common allele (1/1) receiving high cholesterol ( approximately 800 mg/day) diets with different fatty acid compositions. All subjects had the apoE 3/3 genotype. There was no difference in cholesterol absorption between the two genotype groups receiving a high saturated fat diet (33% of total energy as fat; 18% saturated, 3% polyunsaturated, 12% monounsaturated) or a low fat diet (22% of total energy as fat; 7% saturated, 7% polyunsaturated, 8% monounsaturated) diet. However, on a high polyunsaturated fat diet (32% of total energy as fat; 7% saturated, 13% polyunsaturated, 12% monounsaturated) mean fractional cholesterol absorption was 56. 7% +/- 1.9 in 1/1 subjects versus 47.5% +/- 2.1 in 1/2 subjects (P = 0.004). A post hoc analysis of the effect of the apoA-IV T347S polymorphism across all diets revealed a Q360H x T347S interaction on cholesterol absorption, and suggested that the A-IV-2 allele lowers cholesterol only in subjects with the 347 T/T genotype.We conclude that a complex interaction between apoA-IV genotype and dietary fatty acid composition modulates fractional intestinal cholesterol absorption in humans.     

R2R - software to speed the depiction of aesthetic consensus RNA secondary structures

Background  

With continuing identification of novel structured noncoding RNAs, there is an increasing need to create schematic diagrams showing the consensus features of these molecules. RNA structural diagrams are typically made either with general-purpose drawing programs like Adobe Illustrator, or with automated or interactive programs specific to RNA. Unfortunately, the use of applications like Illustrator is extremely time consuming, while existing RNA-specific programs produce figures that are useful, but usually not of the same aesthetic quality as those produced at great cost in Illustrator. Additionally, most existing RNA-specific applications are designed for drawing single RNA molecules, not consensus diagrams.     

An improved method for measuring heart-rate variability: assessment of cardiac autonomic function

Heart rate oscillates in synchrony with respiration. Several methods have been employed to assess this 'sinus arrhythmia', as an index of autonomic nervous system function. This paper proposes a new, easily computed measure, R, which is relatively resistant to the major nonrespiratory sources of variation, including premature beats, heart-rate differences among subjects, and slow trends in heart rate over time within subjects. The method can also be used more generally in any context where individuals are associated with event processes (for example, with seizures), when one requires assessment of the extent to which a point process is periodic with a particular known period.     

Distinctive structure and function of human apolipoprotein variant ApoA-IV-2

We have investigated the molecular structure, phospholipid binding, and lecithin-cholesterol acyltransferase catalytic activity of pure apoA-IV-2, a basic variant isoform of apoA-IV which is inherited as a classical Mendelian allele with a gene frequency of 0.09. Circular dichroism spectroscopy established that the alpha-helical content of apoA-IV-2 was 75% in the native state (versus 56% for apoA-IV-1), and increased to 88% in the presence of phospholipid. Fluorescence titration established that apoA-IV-2 bound to egg phospholipid vesicles with a Ka of 3.3 x 10(6) liter/mol, 2.4-fold greater than the affinity of apoA-IV-1. Fluorescence quenching studies revealed that, unlike apoA-IV-1, binding of apoA-IV-2 to phospholipid vesicles induced strong shielding of the amino-terminal tryptophan against iodide quenching. Enzyme kinetic studies using both saturated and unsaturated phospholipid substrates demonstrated that apoA-IV-2 was 36-71% more efficient in activating lecithin-cholesterol acyltransferase than apoA-IV-1. We conclude that apoA-IV-2 has more alpha-helical structure, is more stable in solution, and is more hydrophobic than apoA-IV-1, and that these distinctive structural features are associated with a higher affinity for phospholipid surfaces and an increased catalytic efficiency of lecithin:cholesterol acyltransferase activation. The biophysical basis for this latter characteristic may be the ability of apoA-IV-2 to penetrate phospholipid surfaces to a greater depth than apoA-IV-1. These molecular properties may be responsible for the increased levels of high density lipoproteins which have been observed in apoA-IV-2 heterozygotes.