Showing papers by "Anthony L. Albiston published in 2007"

Sub-cellular localization of insulin-regulated membrane aminopeptidase, IRAP to vesicles in neurons.

Abstract: Angiotensin IV and LVV-hemorphin 7 promote robust enhancing effects on learning and memory. These peptides are also competitive inhibitors of the insulin-regulated membrane aminopeptidase, suggesting that the biological actions of these peptides may result from inhibition of IRAP activity. However, the normal function of IRAP in the brain is yet to be determined. The present study investigated the sub-cellular distribution of IRAP in four neuronal cell lines and in the mouse brain. Using sub-cellular fractionation, IRAP was found to be enriched in low density microsomes, while lower levels of IRAP were also present in high density microsomes, plasma membrane and mitochondrial fractions. Dual-label immunohistochemistry confirmed the presence of IRAP in vesicles co-localized with the vesicular maker VAMP2, in the trans Golgi network co-localized with TGN 38 and in endosomes co-localized with EEA1. Finally using electron microscopy, IRAP specific immunoreactivity was predominantly associated with large 100–200 nm vesicles in hippocampal neurons. The location, appearance and size of these vesicles are consistent with neurosecretory vesicles. IRAP precipitate was also detected in intracellular structures including the rough endoplasmic reticulum, Golgi stack and mitochondrial membranes. The sub-cellular localization of IRAP in neurons demonstrated in the present study bears striking parallels with distribution of IRAP in insulin responsive cells, where the enzyme plays a role in insulin-regulated glucose uptake. Therefore, we propose that the function of IRAP in neurons may be similar to that in insulin responsive cells.

Insulin-regulated aminopeptidase: analysis of peptide substrate and inhibitor binding to the catalytic domain.

Abstract: Peptide inhibitors of insulin-regulated aminopeptidase (IRAP) accelerate spatial learning and facilitate memory retention and retrieval by binding competitively to the catalytic site of the enzyme and inhibiting its catalytic activity. IRAP belongs to the M1 family of Zn2+-dependent aminopeptidases characterized by a catalytic domain that contains two conserved motifs, the HEXXH(X)18E Zn2+-binding motif and the GXMEN exopeptidase motif. To elucidate the role of GXMEN in binding peptide substrates and competitive inhibitors, site-directed mutagenesis was performed on the motif. Non-conserved mutations of residues G428, A429 and N432 resulted in mutant enzymes with altered catalytic activity, as well as divergent changes in kinetic properties towards the synthetic substrate leucine β-naphthalamide. The affinities of the IRAP inhibitors angiotensin IV, Nle1-angiotensin IV, and LVV-hemorphin-7 were selectively decreased. Substrate degradation studies using the in vitro substrates vasopressin and Leu-enkephalin showed that replacement of G428 by either D, E or Q selectively abolished the catalysis of Leu-enkephalin, while [A429G]IRAP and [N432A]IRAP mutants were incapable of cleaving both substrates. These mutational studies indicate that G428, A429 and N432 are important for binding of both peptide substrates and inhibitors, and confirm previous results demonstrating that peptide IRAP inhibitors competitively bind to its catalytic site.

Angiotensin IV binding site - the AT4 receptor or insulin regulated aminopeptidase (IRAP)

Abstract: Hypertension and Hormonal Mechanisms Chapter Outline (A) New Developments in the Renin-Angiotensin System 1 The Intrarenal Renal-Angiotensin System Minolfa C Prieto-Carrasquero, Hiroyuki Kobori and L Gabriel Navar Tulane University 2 Cardiac and Vascular Renin-Angiotensin Systems Rajesh Kumar, Kenneth M Baker and Jing Pan Texas A & M University Health System 3 Regulation of Cardiovascular Control Mechanisms by Angiotensin (1-7) and Angiotensin Converting Enzyme-2 Carlos M Ferrario, David B Averill, K Bridget Brosnihan, Mark C Chappell, Debra I Diz, Patriciai E Gallagher, Liomar AA Neves and E Ann Tallant Wake Forest University 4 Angiotensin IV and the AT4 Receptor T A Jenkins, FAO Mendelsohn, AL Albiston and SY Chai Howard Florey Institute of Experimental Physiology and Medicine, Melbourne 5 AT2 Receptors in Blood Pressure Regulation Robert M Carey and Helmy M Siragy University of Virginia Health System 6 Angiotensin II and Inflammation Rhian M Touyz and Ernesto L Schiffrin Clinical Research Institute of Montreal 7 Aldosterone and Vascular Damage Hilton V Joffe, Gordon H Williams and Gail K Adler Harvard Medical School (B) The Sympatho-Adrenal System in Hypertension 8 Neurogenic Human Hypertension David Robertson, Andre Diedrich and Italo Biaggioni Vanderbilt University 9 Calcitonin Gene-Related Peptide and Hypertension Donald J DiPette and Scott C Supowit Texas A& M University 10 The Renal Dopaminergic System in Hypertension Pedro A Jose, Robert M Carey and Robin A Felder Georgetown University and the University of Virginia Health System (C) Metabolic Disorders and Hypertension 11 The Insulin Resistance Syndrome and Hypertension James R Sowers University of Missouri 12 Fatty Acids and Hypertension Brent M Egan Medical University of South Carolina 13 Goal-Oriented Hypertension Management in Diabetic and Non-Diabetic Patients Gregory M Singer, John F Setaro and Henry R Black Rush Presbyterian - St Luke's Medical Cente (D) Endothelial Hormones and Autacoids and Hypertension 14 Nitric Oxide and Hypertension David L Mattson and Allen W Cowley Medical College of Wisconsin 15 Role of Endothelin 1 in Hypertension Ernesto L Schiffrin Clinical Research Institute of Montreal 16 The Kallekrein/Kinin System and Hypertension Julie Chao and Lee Chao Medical University of South Carolina (E) Other Hormonal Systems and Hypertension 17 Natriuretic Peptides and Hypertension Kailesh N Pandey Tulane University 18 Sex Steroids and Hypertension Suzanne Oparil and Andrew Miller University of Alabama Birmingham 19 The Lipoxygenase System in the Vasculature and Hypertension Naftali Stern and Michael Tuck Tel Aviv University and UCLA School of Medicine

Angiotensin IV Binding Site

Abstract: Angiotensin IV (Ang IV) is a hexapeptide fragment corresponding to amino acids 3–8 (VYIHPF) of angiotensin II (Ang II) that is formed by consecutive actions of aminopeptidase A and aminopeptidase N (1) (Fig. 1). Ang IV acts as a weak agonist at the Ang II AT1 receptor, and was generally believed to have no physiological role because of its ineffectiveness in the regulation of blood pressure, fluid balance, or adrenal steroid secretion. However, in 1988, specific actions were discovered for Ang IV in the brain—the peptide was found to facilitate memory retention and retrieval (2). A specific, high-affinity binding site was subsequently described in bovine adrenal membranes, which bound Ang IV saturably, reversibly, and with nanomolar affinity (3,4). This binding site was termed as the angiotensin AT4 receptor by an IUPHAR nomenclature committee (5). This AT4 receptor site is pharmacologically distinct from both Ang II AT1 and AT2 receptors, and bound Ang II at only micromolar affinity. Open image in new window Fig. 1. Schematic diagram of the angiotensin system leading to the formation of angiotensin IV.