contain chitin-binding consensus sequence, called the Rebers-Riddiford (R-R)
consensus sequence (Rebers and Riddiford 1988; Rebers and Willis 2001) spanning
about 30 amino acid residues, at the central part of each peptide. The R-R sequence
is found in many cuticle proteins and peptides in arthropods including insects and
crustaceans (Andersen et al. 1995; Andersen 1999; Endo et al. 2000; Faircloth and
Shafer 2007; Ikeya et al. 2001; Shafer et al. 2006; Wynn and Shafer 2005).
However, the similarity is confined only to the R-R sequence, and there is almost
no similarity in the other parts of peptides and proteins of insect cuticle proteins.
Thus, the other part than the R-R sequence may be related to calcification. In
CAP-1, only Ser at the 70th position was phosphorylated among 6 Ser residues.
measurement at 570 nm
22 mM CaCl 2 (100 µl)
22 mM NaHCO 3 (100 µl)
+ sample solution (20 µl)
0
1
2
3
4
5
Time (min)
Absorbance at 570 nm
0
1.2
0.6
every minute
Ca + 2HCO 3
-
CaCO 3 + H 2 CO 3
precipitate
2+
3
-
a
b
Fig. 11.6 Assay method of calcification inhibitory activity.(a) Calcification inhibitory activity of
a sample was assessed by measuring the turbidity (absorbance at 570 nm) of the supersaturated
solution of calcium carbonate. (b) An example of the result. Closed circle: a crude extract of
exoskeleton. Open circle: control
CAP-1
DVDLDEIHQEQNIDDDNTITGSYRWTSPEGVEYFVKYIAD
:.. : . ::
. : : : :.: :: :::.::
CAP-2 SDIIDIEEDHLEHEQEGVPGTAVEGEYSWVAPDGNEYKVKYVAD
RR
G--------G------Y-ACAP-1
EDGYRVLESNAVPATADGVRADGAQGSFVpSSEDDDDDD
:::::: : ::
.
CAP-2
HLGYRVLEDNVVPEVPELEDY
RR
E-GY--------P--P
41
50
60
70
78
1
10
20
30
40
Fig. 11.7 Amino acid sequences of CAP-1 and -2.Acidic amino acid residues are underlined.
Symbols, (:) and (.), between the two sequences represent identical and similar residues, respectively. RR means Rebers-Riddiford consensus sequence for chitin binding
322
H. Nagasawa
consensus sequence (Rebers and Riddiford 1988; Rebers and Willis 2001) spanning
about 30 amino acid residues, at the central part of each peptide. The R-R sequence
is found in many cuticle proteins and peptides in arthropods including insects and
crustaceans (Andersen et al. 1995; Andersen 1999; Endo et al. 2000; Faircloth and
Shafer 2007; Ikeya et al. 2001; Shafer et al. 2006; Wynn and Shafer 2005).
However, the similarity is confined only to the R-R sequence, and there is almost
no similarity in the other parts of peptides and proteins of insect cuticle proteins.
Thus, the other part than the R-R sequence may be related to calcification. In
CAP-1, only Ser at the 70th position was phosphorylated among 6 Ser residues.
measurement at 570 nm
22 mM CaCl 2 (100 µl)
22 mM NaHCO 3 (100 µl)
+ sample solution (20 µl)
0
1
2
3
4
5
Time (min)
Absorbance at 570 nm
0
1.2
0.6
every minute
Ca + 2HCO 3
-
CaCO 3 + H 2 CO 3
precipitate
2+
3
-
a
b
Fig. 11.6 Assay method of calcification inhibitory activity.(a) Calcification inhibitory activity of
a sample was assessed by measuring the turbidity (absorbance at 570 nm) of the supersaturated
solution of calcium carbonate. (b) An example of the result. Closed circle: a crude extract of
exoskeleton. Open circle: control
CAP-1
DVDLDEIHQEQNIDDDNTITGSYRWTSPEGVEYFVKYIAD
:.. : . ::
. : : : :.: :: :::.::
CAP-2 SDIIDIEEDHLEHEQEGVPGTAVEGEYSWVAPDGNEYKVKYVAD
RR
G--------G------Y-ACAP-1
EDGYRVLESNAVPATADGVRADGAQGSFVpSSEDDDDDD
:::::: : ::
.
CAP-2
HLGYRVLEDNVVPEVPELEDY
RR
E-GY--------P--P
41
50
60
70
78
1
10
20
30
40
Fig. 11.7 Amino acid sequences of CAP-1 and -2.Acidic amino acid residues are underlined.
Symbols, (:) and (.), between the two sequences represent identical and similar residues, respectively. RR means Rebers-Riddiford consensus sequence for chitin binding
322
H. Nagasawa