GPCRdb

c5ar1_human

NAME

6C1Q

ORGANISM

Human (Homo sapiens)

PROTEIN NAME

c5ar1_human

SOURCE

SWISSPROT

SEQUENCE

Perspective: Wild-type Construct


N-term
M	D	S	F	N	Y	T	T	P	D
10



Y	G	H	Y	D	D	K	D	T	L
20



D	L	N	T	P	V	D	K	T	S
30


			TM1
N	T	L	R	V	P	D	I	L	A
40



L	V	I	F	A	V	V	F	L	V
50



G	V	L	G	N	A	L	V	V	W
60


					ICL1
V	T	A	F	E	A	K	R	T	I
70


TM2
N	A	I	W	F	L	N	L	A	V
80



A	D	F	L	S	C	L	A	L	P
90



I	L	F	T	S	I	V	Q	H	H
100


ECL1				TM3
H	W	P	F	G	G	A	A	C	S
110



I	L	P	S	L	I	L	L	N	M
120



Y	A	S	I	L	L	L	A	T	I
130



S	A	D	R	F	L	L	V	F	K
140


ICL2
P	I	W	C	Q	N	F	R	G	A
150


TM4
G	L	A	W	I	A	C	A	V	A
160



W	G	L	A	L	L	L	T	I	P
170


					ECL2
S	F	L	Y	R	V	V	R	E	E
180



Y	F	P	P	K	V	L	C	G	V
190


				TM5
D	Y	S	H	D	K	R	R	E	R
200



A	V	A	I	V	R	L	V	L	G
210



F	L	W	P	L	L	T	L	T	I
220



C	Y	T	F	I	L	L	R	T	W
230


		ICL3					TM6
S	R	R	A	T	R	S	T	K	T
240



L	K	V	V	V	A	V	V	A	S
250



F	F	I	F	W	L	P	Y	Q	V
260



T	G	I	M	M	S	F	L	E	P
270


ECL3				TM7
S	S	P	T	F	L	L	L	K	K
280



L	D	S	L	C	V	S	F	A	Y
290



I	N	C	C	I	N	P	I	I	Y
300


				C-term
V	V	A	G	Q	G	F	Q	G	R
310



L	R	K	S	L	P	S	L	L	R
320



N	V	L	T	E	E	S	V	V	R
330



E	S	K	S	F	T	R	S	T	V
340



D	T	M	A	Q	K	T	Q	A	V
350


BRIL (b562R


	N-term						TM1
	S	N	T	L	R	V	P	D	I
38



L	A	L	V	I	F	A	V	V	F
48



L	V	G	V	L	G	N	A	L	V
58



V	W	V	T	A	F	E	A	K	R
68


TM2
T	I	N	A	I	W	F	L	N	L
78



A	V	A	D	F	L	S	C	L	A
88



L	P	I	L	F	T	S	I	V	Q
98


	ECL1					TM3
H	H	H	W	P	F	G	G	A	A
108



C	S	I	L	P	S	L	I	L	L
118



N	M	Y	A	S	I	L	L	L	A
128



T	I	S	A	D	R	F	L	L	V
138


		ICL2
F	K	P	I	W	C	Q	N	F	R
148


TM4
G	A	G	L	A	W	I	A	C	A
158



V	A	W	G	L	A	L	L	L	T
168



I	P	S	F	L	Y	R	V	V	R
178


ECL2
E	E	Y	F	P	P	K	V	L	C
188


						TM5
G	V	D	Y	S	H	D	K	R	R
198



E	R	A	V	A	I	V	R	L	V
208



L	G	F	L	W	P	L	L	T	L
218



T	I	C	Y	T	F	I	L	L	R
228


				ICL3
T	W	S	R	R	A	T	R	S	T
238


TM6
K	T	L	K	V	V	V	A	V	V
248



A	S	F	F	I	F	W	L	P	Y
258



Q	V	T	G	I	M	M	S	F	L
268


ECL3				TM7
E	P	S	S	P	T	F	L	L	L
278



K	K	L	D	S	L	C	V	S	F
288



A	Y	I	N	C	C	I	N	P	I
298



I	Y	V	V	A	G	Q	G	F	Q
308


C-term
G	R	L	R	K	S	L	P	S	L
318



L	R	N	V	L	T	E	E	S	V
328

V

329


TEV proteas



R	E


His(8)

SCHEMATIC

Perspective: Wild-type Construct

N-term

TM1

ICL1

TM2

ECL1

TM3

ICL2

TM4

ECL2

TM5

ICL3

TM6

ECL3

TM7

C-term

BRIL (

N-term

TM1

ICL1

TM2

ECL1

TM3

ICL2

TM4

ECL2

TM5

ICL3

TM6

ECL3

TM7

TEV pr

DIAGRAMS

MUTATIONS

Position	Wild-type	Mutated

DELETIONS

from 1 to 29

from 332 to 350

MODIFICATIONS

AUXILIARY PROTEINS

Protein_type: tag

Name: FLAG

Sequence: None

Deletions:

Positions: N-term_0 None:None

Presence in Crystal:YES

Remarks: None

Protein_type: fusion

Name: BRIL (b562RIL)

Sequence: None

Deletions:

Positions: N-term_0 None:None

Presence in Crystal:YES

Remarks: None

Protein_type: prot_cleavage

Name: TEV protease

Sequence: None

Deletions:

Positions: Within Receptor_0 330:330

Presence in Crystal:YES

Remarks: None

Protein_type: tag

Name: His(8)

Sequence: None

Deletions:

Positions: C-term_0 None:None

Presence in Crystal:NO

Remarks: None

EXPRESSION

Expression method	Host cell type	Host cell	Remarks
Baculovirus	Sf9	Insect	Bac - to - Bac Baculovirus Expression System (Invitrogen)

SOLUBILIZATION

Solvent type	Item	Concentration
detergent	DDM	1 %w/v
additive	CHS	0.2 %w/v

Remarks:

PURIFICATION

Purification type	Description
Proteolysis by TEV protease	None
Detergent DDM exchanged with LMNG	None

Remarks:None

CRYSTALLIZATION

METHOD: in meso/LCP

TYPE: lipidic cubic phase (LCP) (monoolein (9.9 MAG))

PROTEIN CONCENTRATION: 40-50 mg/ml

Temperature: 10°C

ph : 6.5-6.5

Remarks: Crystals grew only at 10°C and not at 20°C

Chemical lists

Type	Item	Concentration
Detergent
	LMNG	0.01 % w/v
Lipid
	CHS	0.001 % w/v
Additional
	PEG 300	25-32 % v/v
	MES, pH 6.5	100.0 mM
	Sodium malonate	90-120 mM
	Polypropylene glycol P400	0.5-1 %