*2.2. Preparation of PPI, CPI, Co, and BL*

Protein samples were prepared using the ISP method [14]. Peas were powdered and sieved through a 100-mesh sieve (0.15 mm). The pea powder was mixed with deionized water at 4 ◦C at a ratio of 1:9 (*w/v*). The pH was adjusted to 10.0 with 1 M NaOH and dissolved for 30 min, followed by centrifugation at 10,000× *g* and 4 ◦C for 20 min. The precipitate was removed and 1 M HCl was slowly added to the supernatant. The mixture was fully stirred, and the pH was adjusted to 5.0. At this time, the precipitation was washed with deionized water, and the pH value was adjusted to 7.0, followed by dialysis. PPI was obtained after freeze-drying for 48 h. The fish muscles were defrosted and chopped, and CPI was prepared according to the steps indicated above for PPI. The pea powder was mixed with fish muscle, and the Co was prepared following the steps described above. BL was obtained by directly mixing the powdered PPI and CPI. To ensure similar comparison, the protein ratio of pea and grass carp in BL and Co was maintained at 1:1 (W:W) in this study.

#### *2.3. Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE)*

Similar concentrations of BL and Co solutions were prepared. Reducing and nonreducing SDS-PAGE gels were used with 5% spacer gel (upper gel) and 12% separation gel (lower gel) on a vertical gel electrophoresis apparatus at 15 mA. The difference between reducing and non-reducing electrophoresis depends on whether or not the reducing agent dithiothreitol (DTT) is added to the loading buffer. Standard protein markers (16–270 kDa) were used for qualitative analysis of the protein bands in the samples. The protein bands were dyed with Coomassie brilliant blue (CBB) R-250. Later, 40% methyl alcohol and 40% acetic acid were used for decoloration. The relative strength of the dyed protein bands was measured using ImageLab (Bio-Rad Laboratories, Hercules, CA, USA).

#### *2.4. Solubility*

According to Kristinsson's method [22], a 0.5% protein solution was prepared. The pH was adjusted to 3.0, 5.0, 7.0, 9.0, and 11.0 and then stirred for 30 min after centrifuging at 10,000× *g* for 20 min. Bovine serum albumin (BSA) was used as the standard and the protein content in the supernatant was measured using the Lowry method at 750 nm. The solubility was calculated as follows:

$$\text{Solubility (\%)} = \text{C/C}\_0 \times 100\%,\tag{1}$$

where *C* is the protein content in the sample after centrifugation (mg) and *C*<sup>0</sup> denotes the protein content in the sample before centrifugation (mg).
