NDLI: The ADAMTS13 metalloprotease domain: roles of subsites in enzyme activity and specificity

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The ADAMTS13 metalloprotease domain: roles of subsites in enzyme activity and specificity

Content Provider	Scilit
Author	Groot, Rens De Lane, David A. Crawley, James T. B.
Copyright Year	2010
Description	ADAMTS13 modulates von Willebrand factor (VWF) platelet-tethering function by proteolysis of the Tyr1605-Met1606 bond in the VWF A2 domain. To examine the role of the metalloprotease domain of ADAMTS13 in scissile bond specificity, we identified 3 variable regions (VR1, -2, and -3) in the ADAMTS family metalloprotease domain that flank the active site, which might be important for specificity. Eight composite sequence swaps (to residues in ADAMTS1 or ADAMTS2) and 18 single-point mutants were generated in these VRs and expressed. Swapping VR1 (E184-R193) of ADAMTS13 with that of ADAMTS1 or ADAMTS2 abolished/severely impaired ADAMTS13 function. Kinetic analysis of VR1 point mutants using VWF115 as a short substrate revealed reduced proteolytic function (kcat/Km reduced by 2- to 10-fold) as a result of D187A, R190A, and R193A substitutions. Analysis of VR2 (F216-V220) revealed a minor importance of this region. Mutants of VR3 (G236-A261) proteolysed wild-type VWF115 normally. However, using either short or full-length VWF substrates containing the P1′ M1606A mutation, we identified residues within VR3 (D252-P256) that influence P1′ amino acid specificity, we hypothesize, by shaping the S1′ pocket. It is concluded that 2 subsites, D187-R193 and D252-P256, in the metalloprotease domain play an important role in cleavage efficiency and site specificity.
Related Links	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2974611/pdf
Ending Page	3072
Page Count	9
Starting Page	3064
DOI	10.1182/blood-2009-12-258780
Journal	Blood
Issue Number	16
Volume Number	116
Language	English
Publisher	American Society of Hematology
Publisher Date	2010-10-21
Access Restriction	Open
Subject Keyword	Biochemistry and Molecular Biology Adamts13 Vwf Metalloprotease Domain Proteolysis Subsites Von Willebrand Factor Kinetic Analysis Vr1 R193 Journal: Blood (Vol- 126, Issue- 16)
Content Type	Text
Resource Type	Article

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