The ovalbumin serpins revisited: Perspective from the chicken genome of clade B serpin evolution in vertebrates -- Benarafa and Remold-O'Donnell 102 (32): 11367 Data Supplement - HTML Page - index.htslp -- Proceedings of the National Academy of Sciences

Benarafa and Remold-O’Donnell. 10.1073/pnas.0502934102.

Supporting Information

Files in this Data Supplement:

Supporting Table 1
Supporting Table 2
Supporting Figure 9
Supporting Figure 10

Supporting Figure 9 Fig. 9.
Amino acid alignment of human and chicken clade B serpins. Residues are colored according to type: polar uncharged (green), acidic (red), basic (blue), and nonpolar (yellow). Secondary structure elements are indicated at the top and are based on the crystal structure of ovalbumin (1OVA) (3). The 60 conserved positions derived from previous studies (7, 8) that were analyzed in all ov-serpins are indicated at the bottom by circles. By convention, numbering refers to that of a1-antitrypsin (7). The interacting residues identified in the atypical human SERPINB5 crystal structure are K¹⁹⁶, T²²⁶, K²⁸³, E³⁴⁹, Q³⁵⁷, and the 5'-stem H³⁵⁸, K³⁵⁹, D³⁶⁰, and E³⁶¹ (32). Blue arrowheads indicate the position of the invariable exon–intron boundaries common for all seven- and eight-exon genes (see Fig. 3C). The orange arrowhead represents the position of the additional boundary found only in eight-exon ov-serpins [shown is the exact position for ovalbumin but varying by a few residues within the CD loop in other eight-exon genes (data not shown)].

Supporting Figure 10

Fig. 10.
Amino acid alignment of human and zebrafish clade B serpins. Residues are colored according to type: polar uncharged (green), acidic (red), basic (blue), and nonpolar (yellow). Secondary structure elements are indicated at the top and are based on the crystal structure of ovalbumin (1OVA) (3). The 60 conserved positions derived from previous studies (7, 8) that were analyzed in all ov-serpins are indicated at the bottom by circles. By convention, numbering refers to that of a1-antitrypsin (7). The interacting residues identified in the atypical human SERPINB5 crystal structure are K¹⁹⁶, T²²⁶, K²⁸³, E³⁴⁹, Q³⁵⁷, and the 5'-stem H³⁵⁸, K³⁵⁹, D³⁶⁰, and E³⁶¹ (32).

Table 1. Accession numbers of chicken clade B serpins

Build 1.1 locus name	Common name	Proposed official serpin nomenclature	Accession nos.	Current annotation
LOC420894		serpinb1	XP_418980	Similar to serine (or cysteine) proteinase inhibitor, clade B (ovalbumin), member 1
LOC420895		serpinb6	XP_418981	Similar to MGC64421 protein
LOC428483		serpinb10	XP_426040*	Similar to heterochromatin-associated protein MENT
MENT-1	MENT	serpinb10b	NP_990228	Heterochromatin-associated protein MENT
LOC420896		serpinb2	XP_418982	Similar to plasminogen activator inhibitor
LOC396058	Ovalbumin	serpinb14	NP_990483	Ovalbumin
LOC420897	Gene Y	serpinb14b	XP_418983	Similar to ovalbumin-related Y protein, chicken
LOC420898	Gene X	serpinb14c	XP_418984^†	Similar to ovalbumin-related Y protein, chicken
LOC420899		serpinb12	XP_418985	Similar to serine (or cysteine) proteinase inhibitor, clade B (ovalbumin), member 12
LOC420900		serpinb5	XP_418986^‡	Similar to maspin

The proposed nomenclature for the chicken clade B serpins follows the rules of the Serpin Gene Nomenclature Committee (2).

*XP_426040 is a predicted sequence not found in EST databases that lacks one exon. By using the corresponding sequence of MENT, the missing exon was found in intronic sequence. The exon was also found by using genscan (score of 0.08) and begins at position 1362 and ends at 1448 in the genomic sequence and encodes for 29 amino acids with compatible phasing with adjacent exons.

^†The first 108 amino acids of predicted sequence XP_418984 were excluded from our study. The position of Met109 is identical to that of Met1 of ovalbumin and gene Y and is at the conserved position for translation start site in exon 2. ^‡The first 283 amino acids of predicted XP_418986 were excluded from our study. The position of Met284 is identical to Met1 of mammalian SERPINB5 (maspin) and the ATG encoding it is in the conserved position in exon 2.

Table 2. Ov-serpin genes in zebrafish (Danio rerio)

Zebrafish	Human						Chicken						Zebrafish						Gene structure	P1–P1'	Accession nos.
Zebrafish	B1	B5	B6	B2	B10	B12	b1	b5	b6	b2	b10	b12	ZGC:91981	CAI20745	ZGC:64178	AAQ97848	ZGC:77645	ZGC:76926	Gene structure	P1–P1'	Protein	Nucleotide
ZGC:91981	57	36	48	43	43	40	56	35	53	45	43	43	x	75	47	51	46	36	Seven-exon	Cys–Met	ZGC:91981	BC076524
CAI20745	51	35	45	41	44	39	52	36	52	46	42	41	75	x	46	51	45	34	Seven-exon	Ser–Ala	CAI20745	BX248515
ZGC:64178	49	36	51	39	40	41	50	37	52	42	42	40	47	46	x	55	48	34	Seven-exon	Arg–Cys	ZGC:64178	BC053300
AAQ97848	58	39	50	44	47	46	56	39	55	47	49	44	51	51	55	x	75	38	Seven-exon	Leu–Cys	AAQ97848	AY398415
ZGC:77645*	47	34	46	44	45	43	46	35	48	47	46	44	46	45	48	75	x	33	Eight-exon	Arg–Thr	ZGC:77645	BC064292
ZGC:76926	34	28	34	32	30	29	33	30	35	32	32	31	36	34	34	38	33	x	n/a	Leu–Cys	ZGC:76926	BC066740

The names for the zebrafish ov-serpins starting with ZGC were characterized by full-length cDNAs isolated as part of the zebrafish gene collection (ZGC). The remaining ov-serpins are designated by their protein accession nos. CAI20745 is a predicted sequence from genomic scaffold BX248515. This locus also includes ZGC:91981 (under protein accession no. CAI20749) and a serpinb1-like pseudogene CAI20747, not shown in the table (when the premature stop codon is omitted, the predicted protein sequence of CAI20747 is 76% and 80% identical to ZGC:91981 and CAI20745, respectively). Inter- and intraspecies percentage of protein sequence identity is shown (x indicates intersection of the same zebrafish protein, i.e. 100% identity). The gene-structure information was not available (n/a) for ZGC:76926.

*The predicted coding sequence of ZGC:77645, currently in the database, is based on the mRNA transcript. A more likely start codon was identified by genscan 120 bp upstream, adding 40 amino acids at the N terminus, and that start codon was used in this study.