Current Search: Phospholipids (x)
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- Title
- ROLE OF MEMBRANE LIPIDS IN MODULATING PROTEIN STRUCTURE & FUNCTION.
- Creator
-
Ray, Supriyo, Tatulian, Suren, University of Central Florida
- Abstract / Description
-
A-B family of toxins consists of plant toxins such as ricin and bacterial toxins such as cholera. The A subunit is the enzymatic domain and the B subunit is the receptor binding domain. Commonly, these toxins bind to the target cell plasma membrane receptors through their B subunit followed by endocytosis and a transport to the endoplasmic reticulum (ER). Inside the ER, the A subunit dissociates from the rest of the toxin, unfolds and triggers the ER quality control mechanism of ER-associated...
Show moreA-B family of toxins consists of plant toxins such as ricin and bacterial toxins such as cholera. The A subunit is the enzymatic domain and the B subunit is the receptor binding domain. Commonly, these toxins bind to the target cell plasma membrane receptors through their B subunit followed by endocytosis and a transport to the endoplasmic reticulum (ER). Inside the ER, the A subunit dissociates from the rest of the toxin, unfolds and triggers the ER quality control mechanism of ER-associated degradation (ERAD). Most ERAD substrates are purged out of the ER into the cytosol for proteasomal degradation. However, the low content of lysine amino acid residues allows the toxin to evade polyubiquitination and subsequent proteasomal degradation. The toxin A subunit refolds into an active conformation in the cytosol, setting off downstream toxic events. In the first part of my thesis, the hypothesis was tested that inhibiting the unfolding of the toxin A subunit inside the ER will prevent ERAD activation, toxin export to the cytosol and intoxication. The chemical chaperones glycerol and sodium 4-phenyl butyrate (PBA) were used to inhibit the toxin A chain unfolding. In vitro biophysical experiments indicated that both chemical chaperones indeed stabilize the cholera toxin A subunit and prevent cytotoxicity. In case of ricin, both chaperones stabilized the toxin A chain but only glycerol prevented cytotoxicity. Additional experiments showed that PBA-treated ricin A chain is destabilized when exposed to anionic lipid membranes mimicking the properties of the ER membrane. In contrast, anionic lipid did not prevent ricin A chain stabilization by glycerol. This explains why glycerol but not PBA blocked ricin intoxication, as only glycerol stabilizes ricin A chain in the presence of ER membranes. Cholera toxin in contrast, remained either unaffected or slightly stabilized in presence of anionic lipids both in presence and absence of PBA. This shows that destabilization by anionic lipids is a toxin-specific rather than a general effect. In the second part of my thesis, the effect of inner leaflet of plasma membrane on the structure of cholera toxin A chain (CTA1) was studied. Since CTA1 refolds into an active conformation in the cytosol in association with unidentified host factors, I hypothesized that inner leaflet of the plasma membrane might play a role to stabilization and/or refolding of CTA1. CTA1 was shown to be a membrane interacting protein, and membranes mimicking lipid rafts had a significant stabilizing effect on its structure. Lipid rafts helped in the regaining of the tertiary and secondary structure of CTA1, while non-raft lipids had a smaller stabilizing effect on CTA1 structure. In the next part of my thesis, I studied the effect of membrane binding on the structure and function of human pancreatic phospholipase A2 (PLA2). Lipid thermal phase transition was found to have a dramatic effect on PLA2 activity. It was also established that although membrane binding and insertion was essential for of PLA2 activity, lipid structural heterogeneity was more important than the depth of membrane insertion for enzyme activation. Most importantly, significant changes in PLA2 secondary and tertiary structures were identified that evidently contribute to the interfacial activation of PLA2. Overall, we conclude that the function of membrane binding enzymes can be significantly modulated via conformational changes induced by interactions with membranes. Thus, we have elucidated various roles of membrane lipids from unfolding and refolding to activation and modulation of membrane binding enzymes. Physical properties of lipids help in regulating various aspects of protein structure and function and their analysis helped us in appreciating the influence wielded by the membrane lipids in the enzyme's surrounding environment.
Show less - Date Issued
- 2011
- Identifier
- CFE0004035, ucf:49184
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0004035
- Title
- The Effects of Phosphatidylserine on Reaction Time and Cognitive Function Following an Exercise Stress.
- Creator
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Wells, Adam, Hoffman, Jay, Fragala, Maren, Stout, Jeffrey, University of Central Florida
- Abstract / Description
-
Phosphatidylserine (PS) is an endogenously occurring phospholipid that has been shown to have cognition and mood enhancing properties in humans, possibly through its role as an enzyme co-factor in cellular signal transduction. Specifically, PS has been identified as activator of classical isoforms of protein kinase C, an enzyme known to be involved in the growth and differentiation of neural cells, and is therefore thought to play a role in the protection of neurons.The purpose of this study...
Show morePhosphatidylserine (PS) is an endogenously occurring phospholipid that has been shown to have cognition and mood enhancing properties in humans, possibly through its role as an enzyme co-factor in cellular signal transduction. Specifically, PS has been identified as activator of classical isoforms of protein kinase C, an enzyme known to be involved in the growth and differentiation of neural cells, and is therefore thought to play a role in the protection of neurons.The purpose of this study was to examine the effects of supplementation with PS and caffeine on measures of cognition, reaction time and mood prior to and following an exercise stress. Twenty, healthy, resistance trained males (17) and females (3) (mean (&)#177; SD; age: 22.75 (&)#177; 3.27 yrs; height: 177.03 (&)#177; 8.44cm; weight: 78.98 (&)#177; 11.24kg; body fat%: 14.28 (&)#177; 6.6), volunteered to participate in this randomized, double-blind, placebo-controlled study. Participants were assigned to a PS group (400mg/day PS; 100mg/day caffeine, N=9) or PL (16g/day Carbs, N=11) delivered in the form of 4 candy chews identical in size, shape and color. Subjects performed an acute bout of full body resistance exercise, prior to (T1) and following 14 days of supplementation (T2). Measures of reaction time (Dynavision(&)#174; D2 Visuomotor Training Device), cognition (Serial Subtraction Test, SST), and mood (Profile of Mood States, POMS) were assessed immediately before and following resistance exercise in both T1 and T2. Data was analyzed using two-way ANCOVA and repeated measures ANOVA.Supplementation with 400mg PS and 100mg caffeine did not have a significant impact upon measures of reaction time or cognition between groups at baseline or following acute resistance exercise. However, there was a non-significant trend to the attenuation of fatigue between groups, following acute resistance exercise (p = 0.071). Interestingly, our data suggests that acute resistance exercise alone may improve cognitive function.Although more research is necessary regarding optimal dosage and supplementation duration, the current findings suggest that supplementation 400mg/day PS with 100mg/day caffeine may attenuate fatigue following acute resistance exercise. It is possible that the lack of significance may be the result of both an inhibition of the PS activated pathway and a withdrawal effect from caffeine.
Show less - Date Issued
- 2012
- Identifier
- CFE0004457, ucf:49325
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0004457
