Table 2.

Apparent in vitro dephosphorylation rates of NHE1 by CN

NHE1ct or CN variant	Site	Phosphosite sequencea	kdephos (10−3 h−1)^
NHE1ct WT and variantsb
NHE1ct
NHE1ct WT	pT779	771SPGTDDVFTPAPSDSPS787	98 ± 3
NHE1ct motif variants
NHE1ctAVATAA	pT779	771SPGTDDVFTPAPSDSPS787	11 ± 3
NHE1ctATAP	pT779	771SPGTDDVFTPAPSDSPS787	642 ± 28
NHE1ctAVATAA/ATAP	pT779	771SPGTDDVFTPAPSDSPS787	811 ± 55
NHE1ct dynamic linker variant
NHE1ctRARAc	pT779	771SPGTDDVFTPAPSDSPS787	199 ± 10
Role of Thr vs. Ser in CN-mediated dephosphorylation
NHE1ct T779S	pT779S	771SPGTDDVFSPAPSDSPS787	1 ± 2
NHE1ct S785T	pT779 pS785T	771SPGTDDVFTPAPSDTPS787     777VFTPAPSDTPSSQRIQ792	193 ± 5 35 ± 7
Role of TxxP motif in CN-mediated dephosphorylation
NHE1ct P782A	pT779	771SPGTDDVFTPAASDSPS787	28 ± 3
NHE1ct S785T/S787A/S788P      785TPAP788	pT779 pS785T	771SPGTDDVFTPAPSDTPA787     777VFTPAPSDTPAPQRIQ792	102 ± 7 96 ± 7
CN variantsd
CNA	pT779	771SPGTDDVFTPAPSDSPS787	347 ± 10
CNAAc	pT779	771SPGTDDVFTPAPSDSPS787	60 ± 10

^aSequence: dephosphorylated residue (underlined)

^bAll reactions performed using CN and the NHE1ct constructs indicated

^cReactions performed with a 20:1 (vs.100:1) NHE1ct:CN ratio and the rates normalized using wt-NHE1ct

^dAll reactions performed using NHE1ct with the CN constructs indicated

^Apparent rates of dephosphorylation were extracted from global non-linear least-square fits of disappearing peaks (dephosphorylation) to single exponentials in SigmaPlot. The standard errors (±) of the estimated parameters were reported to represent confidence intervals