225 research outputs found
Crystal structure of peptide-bound neprilysin reveals key binding interactions
Neprilysin (NEP) is a promiscuous zinc metalloprotease with broad substrate specificity and cleaves a remarkable diversity of substrates through endopeptidase action. Two of these – amyloid-β and natriuretic peptides – implicate the enzyme in both Alzheimer’s disease and cardiovascular disease, respectively. Here, we report the creation of a catalytically inactive NEP (E584D) to determine the first peptide-bound crystal structure at 2.6 Å resolution. The structure reveals key interactions involved in substrate binding which we have identified to be conserved in other known zinc metalloproteases. In addition, the structure provides evidence for a potential exosite within the central cavity that may play a critical role in substrate positioning. Together, these results contribute to our understanding of the molecular function of NEP.</p
Crystal structure of peptide-bound neprilysin reveals key binding interactions
Neprilysin (NEP) is a promiscuous zinc metalloprotease with broad substrate specificity and cleaves a remarkable diversity of substrates through endopeptidase action. Two of these – amyloid-β and natriuretic peptides – implicate the enzyme in both Alzheimer’s disease and cardiovascular disease, respectively. Here, we report the creation of a catalytically inactive NEP (E584D) to determine the first peptide-bound crystal structure at 2.6 Å resolution. The structure reveals key interactions involved in substrate binding which we have identified to be conserved in other known zinc metalloproteases. In addition, the structure provides evidence for a potential exosite within the central cavity that may play a critical role in substrate positioning. Together, these results contribute to our understanding of the molecular function of NEP.</p
Structure of mouse IP-10, a chemokine
The structure of mouse IP-10 shows a novel tetrameric association
C9orf72, a protein associated with amyotrophic lateral sclerosis (ALS) is a guanine nucleotide exchange factor
Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two late onset neurodegenerative diseases, have been shown to share overlapping cellular pathologies and genetic origins. Studies suggest that a hexanucleotide repeat expansion in the first intron of the C9orf72 gene is the most common cause of familial FTD and ALS pathology. The C9orf72 protein is predicted to be a differentially expressed in normal and neoplastic cells domain protein implying that C9orf72 functions as a guanine nucleotide exchange factor (GEF) to regulate specific Rab GTPases. Reported studies thus far point to a putative role for C9orf72 in lysosome biogenesis, vesicular trafficking, autophagy and mechanistic target of rapamycin complex1 (mTORC1) signaling. Here we report the expression, purification and biochemical characterization of C9orf72 protein. We conclusively show that C9orf72 is a GEF. The distinctive presence of both Rab- and Rho-GTPase GEF activities suggests that C9orf72 may function as a dual exchange factor coupling physiological functions such as cytoskeleton modulation and autophagy with endocytosis
High resolution crystal structure of substrate-free human neprilysin
Neprilysin is a transmembrane M13 zinc metalloprotease responsible for the degradation of several biologically active peptides including insulin, enkephalin, substance P, bradykinin, endothelin-1, neurotensin and amyloid-β. The protein has received attention for its role in modulating blood pressure responses with its inhibition producing an antihypertensive response. To date, several inhibitor bound crystal structures of the human neprilysin extracellular domain have been determined, but, a structure free of bound inhibitor or substrate has yet to be reported. Here, we report the first crystal structure free of substrate or inhibitor for the extracellular catalytic domain of human neprilysin at 1.9 Å resolution. This structure will provide a reference point for comparisons to future inhibitor or substrate bound structures. The neprilysin structure also reveals that a closed protein conformation can be adopted in protein crystals absent of bound substrate or inhibitor.</p
The Design and Development of a Potent and Selective Novel Diprolyl Derivative That Binds to the N-Domain of Angiotensin-I Converting Enzyme
Angiotensin-I converting enzyme (ACE)
is a zinc metalloÂprotease
consisting of two catalytic domains (N- and C-). Most clinical ACE
inhibitor(s) (ACEi) have been shown to inhibit both domains nonselectively,
resulting in adverse effects such as cough and angioedema. Selectively
inhibiting the individual domains is likely to reduce these effects
and potentially treat fibrosis in addition to hypertension. ACEi from
the GVK Biosciences database were inspected for possible N-domain
selective binding patterns. From this set, a diprolyl chemical series
was modeled using docking simulations. The series was expanded based
on key target interactions involving residues known to impart N-domain
selectivity. In total, seven diprolyl compounds were synthesized and
tested for N-domain selective ACE inhibition. One compound with an
aspartic acid in the P<sub>2</sub> position (compound <b>16</b>) displayed potent inhibition (<i>K</i><sub>i</sub> = 11.45
nM) and was 84-fold more selective toward the N-domain. A high-resolution
crystal structure of compound <b>16</b> in complex with the
N-domain revealed the molecular basis for the observed selectivity
Identification of Placenta Growth Factor Determinants for Binding and Activation of Flt-1 Receptor
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